Ophthalmic composition useful in particular for the administration of active principles that are sparingly soluble in water or relatively unstable in water

Abstract

The present invention relates to an ophthalmic composition, comprising a nonionic oil-in-water emulsion with: • an aqueous phase Aq, • a lipid phase L, in the form of droplets dispersed in the aqueous phase, • an interface between said lipid phase L and said aqueous phase Aq, and • a particulate surface-active agent for stabilizing the emulsion, located at the interface between the lipid phase and the aqueous phase, said surface-active agent comprising or consisting of organic particles with an average size of less than or equal to 1 μm, said composition being free of non-particulate surfactant, and advantageously free of preservative. These compositions are useful as a medicament, in particular for administration by ocular instillation. The invention also relates to a method for preparing said compositions.

Description

[0001] DESCRIPTION

[0002] Ophthalmic composition useful in particular for the administration of active ingredients which are not very water-soluble or not very stable in water

[0003] FIELD OF THE INVENTION

[0004] The invention relates to ophthalmic compositions comprising a non-ionic oil-in-water emulsion, and optionally a poorly water-soluble active ingredient, of BCS class II or IV.

[0005] STATE OF THE ART

[0006] The aim of administering active substances via the ophthalmic route is to achieve a therapeutic concentration and maintain it over time. However, the various protective mechanisms of the eye run counter to these treatment objectives.

[0007] In the case of ophthalmic administration, the first challenge is to be able to maintain a sufficiently long residence time at the administration site to ensure absorption of the active ingredient. Indeed, blinking of the eyelids and the significant clearance of the tear film contribute to rapid evacuation of administered liquids. During blinking, the eyelid allows better spreading of the tear film on the corneal surface, but also promotes its flow. The volume of tear fluid, approximately 7 L, is completely renewed every 5 minutes, leading to rapid elimination of the administered active ingredients.

[0008] Thus, an isotonic formula with a pH close to that of the tear fluid (pH of approximately 7.4) will be better tolerated on the ocular surface and less likely to be eliminated by tears. Similarly, a viscous formula will be eliminated more slowly. Although diluted in the tear film upon administration, the formulation can therefore play a role in tear clearance.

[0009] The second challenge is to ensure good ocular bioavailability of the active ingredient. While increasing the residence time on the surface of the eye extends the window of time available for absorption of the active ingredient, it is not enough. The active ingredient still has to cross the various barriers.

[0010] In the case of anterior eye pathologies, it is the cornea. The cornea is composed of 5 layers:

[0011] 1. the corneal epithelium, which is a lipid-rich, non-keratinized stratified squamous epithelium representing approximately 10% of the corneal thickness. The cells are bound by tight junctions, conferring the role of a barrier to the diffusion of the epithelium;

[0012] 2. Bowman's layer, located under the basal lamina of the epithelium, is a dense layer derived from the stroma consisting of collagen fibers with a random organization; 3. the stroma, which constitutes almost 90% of the thickness of the cornea. It is made up of keratocytes producing collagen fibers with a regular orientation and the ground substance rich in water and glycoproteins;

[0013] 4. Descement's membrane, which is the basal layer of the endothelium, rich in type IV collagen fibers and whose thickness increases with age;

[0014] 5. Finally, the endothelium, composed of a monolayer of hexagonal cells derived from neural crests. Active ion transport by the endothelium allows the transfer of water from the stroma and maintains the transparency of the stroma.

[0015] All layers of the cornea, due to their different structures and properties, therefore act as barriers to the absorption of active ingredients. Thus, it has been estimated that when administering a conventional ophthalmic solution, less than 10% of the active ingredient is absorbed.

[0016] In addition to these issues common to all active ingredients, the formulation of active ingredients that are poorly soluble or unstable in water poses particular challenges. To illustrate this, we will take the example of formulations based on prostaglandins or their analogues (PGA), useful in the treatment of glaucoma, and more particularly in the treatment of open-angle glaucoma. Indeed, it has been reported that approximately 25% of patients were not compliant with their treatment (non-compliance defined as the cancellation of at least one administration per week or the inability of the patient to accurately describe their dosing schedule), mainly due to forgetfulness but also to the occurrence of adverse effects.

[0017] Local adverse effects restrict the use of PGAs even though their hypotensive effect has been shown to be the most important among antiglaucoma treatments.

[0018] This may be explained by the fact that in the relatively wide range of pharmaceutical formulations of PGA, most contain preservatives and / or solubilizing agents such as benzalkonium chloride (BAK), or non-particulate surfactants such as macrogolglycerol hydroxystearate (MGHS).

[0019] However, the use of preservatives, particularly BAK, in eye drops is problematic. Numerous studies comparing formulas with BAK versus preservative-free formulas have been conducted, whether for latanoprost, travoprost, or bimatoprost. All have concluded that BAK has a harmful effect.

[0020] Furthermore, most preservative-free PGA formulations use MGHS as a solubilizing and stabilizing agent. However, the in vitro cytotoxicity of MGHS has recently been highlighted. MGHS has also been shown in vitro to significantly induce necrosis and activation of the P2X7 receptor, a receptor linked to various cellular effects ranging from inflammation and oxidative stress to apoptosis. Furthermore, studies report the deleterious effects of MGHS, among other surfactants, on the ocular surface. Therefore, there is still a need for formulations for molecules in this pharmacological class.

[0021] Faced with this problem, alternative aqueous formulations have been developed. These include:

[0022] • surfactant-based formulations, such as micellar solutions, liposomes and niosomes. Examples of such formulations in which non-ionic surfactants (PS80, also noted Tween 80) make it possible to solubilize a prostaglandin derivative are described in particular in Alviset et al. Pharmaceutics 2022, 14, 453;

[0023] • formulations based on cyclodextrins;

[0024] • hydrogel-based formulations;

[0025] • formulations based on polymeric nanoparticles.

[0026] Oil-in-water (O / W) emulsions are also common for solubilizing and stabilizing poorly soluble or unstable active ingredients in water. With a sufficiently high partition coefficient of the active ingredient between oil and water, solubilizing a sensitive active ingredient in oil droplets can protect it. This phenomenon has been demonstrated by several teams for latanoprost in particular.

[0027] Another advantage of O / W emulsions, specific to ocular administration, is the interaction with the tear film. One example is Santen pharmaceutical's Novasorb® technology, based on eye drops comprising a cationic O / W emulsion. The emulsion's positive charges allow for close interaction with the negatively charged mucin layer covering the cornea, while the lipophilic part of the emulsion can spread over the surface and simulate the lipid layer of the tear film. This mechanical action has led to the commercialization of Cationorm®, artificial tears for treating dry eye. DE-130A, or Catioprost®, a preservative-free cationic emulsion containing latanoprost, for treating glaucoma while protecting the ocular surface, is currently under development (phase III trials began in 2019 and a marketing authorization application has been filed).

[0028] International application WO2021 / 113637 also describes cationic emulsions, intended for ophthalmic administration, in particular for the treatment of presbyopia. In these cationic emulsions, it is noted that the active ingredient can be coated with various substances in order to modify its release kinetics. These emulsions can contain nanoparticles, but only silica is given as an example of nanoparticles. However, silica is irritating to the cornea and non-biodegradable, which does not seem compatible with ocular administration. In addition, the use of benzalkonium chloride as a cationic agent is envisaged. O / W nanoemulsions of travoprost at 1% w / v oil phase, stabilized with PS80 at 2% w / w, have also been described (Ismail et al. Int. J. Pharm. 2020, 583, 119402). The ocular bioavailability of travoprost was increased twofold with this formulation.This effect has been attributed to the small size of the nano-droplets and their oily nature increasing their residence time on the corneal surface as well as to the presence of PS80 which can act as an absorption promoter. Nevertheless, the stability of the product (three months at 25°C under a relative humidity of 60%) has proven to be largely insufficient for storage at room temperature. Thus, a need remains for new aqueous formulations, essentially free of preservatives irritating to the cornea, particularly during repeated applications, and avoiding the use of solubilizers or non-particulate surfactants - even non-ionic ones - while remaining stable at room temperature over periods of several months. Optionally, the formulations include an active ingredient, particularly one which is not soluble or not very stable in water, and improve its stability and / or bioavailability during ocular administration.

[0029] BRIEF STATEMENT OF THE INVENTION

[0030] To address this, the inventors propose new ophthalmic compositions, comprising a Pickering emulsion (also called Ramsden) stabilized using organic particles with specific properties.

[0031] Thus, according to a first aspect, the invention relates to an ophthalmic composition, comprising an oil-in-water emulsion with:

[0032] • an aqueous phase Aq,

[0033] • a lipid phase L, in the form of droplets dispersed in the aqueous phase,

[0034] • an interface between said lipid phase L and said aqueous phase Aq, and

[0035] • a particulate surfactant for stabilizing the emulsion, located at the interface between the lipid phase and the aqueous phase, said surfactant comprising or consisting of organic particles of average size (in intensity) less than or equal to 1 pm, preferably less than or equal to 500 nm, said composition being free of non-particulate surfactant, and preferably free of preservative, said lipid phase L optionally comprising at least one liposoluble active ingredient.

[0036] The ophthalmic compositions of the invention are stable at room temperature, and this for periods of several months.

[0037] The ophthalmic compositions of the invention make it possible to dispense with the use of (non-particulate) surfactants. Thus, they also have improved tolerance compared to the compositions of the prior art, particularly when they must be administered repeatedly. In addition, the lipid globules help to reconstitute the lubricating lipid films protecting the eye, which improves tolerance, and extends the residence time of the active ingredient on the eye.

[0038] Thus, where appropriate, the ophthalmic compositions of the invention also improve the adsorption of the active ingredient because the lipid globules disintegrate rapidly upon contact with the eye and biological fluids.

[0039] According to another aspect, the invention relates to said composition for its use as a medicament, in particular for administration by ocular instillation.

[0040] According to another aspect, the invention relates to a method of preparing the emulsion.

[0041] According to a particular aspect of the invention, the active ingredient is chosen from prostaglandins. These particular compositions are useful as medicaments, in particular in the treatment or prevention of glaucoma, such as open-angle glaucoma.

[0042] According to another particular aspect of the invention, the active ingredient is chosen from steroids. These particular compositions are useful as a medicament, in particular for preventing, relieving or treating ocular inflammations.

[0043] DETAILED DESCRIPTION OF THE INVENTION

[0044] Ophthalmic compositions

[0045] In the present invention, the term "pharmaceutical acceptable" is intended to mean that which is generally safe, non-toxic, neither biologically nor otherwise undesirable, and which is acceptable for veterinary or human pharmaceutical use, in particular by ocular administration.

[0046] As used herein, the term "pharmaceutical composition" is intended to mean a composition which is generally safe, non-toxic and neither biologically nor otherwise undesirable, and which is acceptable for veterinary or human pharmaceutical use, particularly for ocular administration.

[0047] “Ocular administration” means topical application to the eye. Thus, the composition can be applied to the eye and more particularly to the cornea, in particular by ocular instillation. Instillation can take place in different sites of the eye, more particularly at the level of the cornea, the conjunctiva, and in particular at the level of the conjunctival sac.

[0048] For the purposes of the present invention, the term "ophthalmic composition" means a pharmaceutical composition for ocular administration, such as eye drops. Such compositions are formulated so as to respect: good tolerance (in particular acceptable pH) and physiological osmolarity. It is furthermore preferable that the compositions are sterile and formulated so as to avoid bacterial contamination during use. As galenic formulations for ocular administration, mention may in particular be made of: • Eye drops: in the form of a sterile emulsion, containing one or more active ingredients. The eye drops can be packaged in multi-dose or single-dose packaging.

[0049] • Eye ointments, which are semi-solid, sterile preparations intended to be applied to the conjunctiva, containing one or more active ingredients and suitable excipients (petroleum jelly, liquid paraffin).

[0050] • Ophthalmic gels, i.e. sterile, semi-solid preparations intended to be applied to the conjunctiva, containing one or more active ingredients and suitable excipients. The excipient is advantageously a hydrophilic polymer which gels in the presence of water, such as hyaluronic acid or a hyaluronate salt such as sodium hyaluronate, a cellulose derivative, a gellan gum, a xanthan gum, or a synthetic polymer such as a carbomer, carbopol®, polyacrylic acid or one of its derivatives. The cellulose derivative is preferably chosen from hydroxyethylcellulose (for example NATROSOL 250M®, NATROSOL 250G®) and / or hydroxypropylmethylcellulose.

[0051] • Ophthalmic inserts, i.e. sterile solid or semi-solid preparations, intended to be inserted into the conjunctival sac. They generally consist of a reservoir of active ingredient embedded in a matrix surrounded by a membrane to control the release. The active ingredient is released gradually.

[0052] Eye drops are preferred because it is easier for a patient to administer such a composition by instilling 1 or 2 drops of the solution, suspension, emulsion, or semi-solid gel into the affected eye. However, the composition may also be a suspension, a viscous or semi-viscous gel, or other types of solid or semi-solid compositions.

[0053] The ophthalmic composition is free of non-particulate surfactants. Such compositions in fact have better tolerance on the ophthalmic mucosa, because non-particulate surfactants (particularly ionic) generate problems of cellular toxicity as indicated above. Surfactants are well known to those skilled in the art: they are a molecule or a salt capable of modifying the surface tension of the emulsion, particularly at the interface between the lipid L and aqueous Aq phases. Non-particulate surfactants are distinguished from particulate surfactants in that they do not comprise particles, whether solid or semi-solid. There are three categories of non-particulate surfactants: anionic surfactants, cationic surfactants, and non-ionic surfactants.

[0054] For the purposes of the invention, a “composition free of non-particulate surfactants” means a composition comprising approximately 0% by weight, relative to the total weight of the composition, of surfactant, whether ionic or non-ionic. In other words, no surfactant is added to the composition. Indeed, even non-ionic surfactants are problematic for ocular tolerance, because just like ionic surfactants, they destabilize biological membranes. Their use must therefore be particularly limited in ophthalmic compositions, particularly in terms of concentrations.The composition is in particular free of surfactants chosen from the group consisting of BAK, MGHS 15, MGHS 35, MGHS 40, glycerol monostearate, hydrogenated polyoxyethylene stearate, PEG-PPG copolymers, nonoxynol-9, octoxynol-40, poloxamer 188, poloxamer 407, polysorbate 20, polysorbate 80, polyoxylethylene stearate (in particular 40), tyloxapol, and mixtures thereof.

[0055] Preferably, the ophthalmic composition is sterile, that is to say that it is devoid or essentially devoid of any form of life, such as in particular bacteria or microorganisms such as yeast and mold.

[0056] Advantageously, the ophthalmic composition is free of preservatives with antimicrobial or microstatic properties, and preferably free of cyclodextrin.

[0057] As used herein, a "preservative" or "preservative agent" has microbicidal or microstatic properties in the composition, i.e., a microbicidal agent is an agent that kills microbes, while a microstatic agent only inhibits the growth of microbes. Preservatives with antimicrobial ( / .e., microbicidal or microstatic) properties should be distinguished from auxiliary agents involved in maintaining the physicochemical characteristics of the active compound or solution: examples of such auxiliary agents include, but are not limited to, EDTA and vitamin E. Furthermore, some preservatives have surface-active properties.

[0058] The ophthalmic composition is advantageously free of surfactant preservatives. Certain preservatives (non-surfactants) may nevertheless be present in the composition of the invention.

[0059] Preferably, the composition is therefore free of surfactant preservatives, in particular auxiliary agents involved in maintaining the physicochemical characteristics of the active compound or the solution. Thus, the composition will advantageously comprise 0% of auxiliary agent such as: sorbic acid and its salts, and quaternary ammonium halides, in particular quaternary ammonium chloride and bromide such as, for example, BAK, benzododecinium halides (such as, for example, benzododecinium bromide), benzethonium halides (such as, for example, benzethonium chloride), lauralkonium halides (such as, for example, lauralkonium chloride), cetrimide, hexadecyltrimethylammonium halide halides (such as, for example, hexadecyltrimethylammonium bromide), tetradecyltrimethylammonium halides (such as, for example, tetradecyltrimethylammonium bromide),dodecyltrimethylammonium halides (such as, for example, dodecyltrimethylammonium bromide), cetrimonium halides, behenalkonium halides, cetalkonium halides (such as, for example, cetalkonium chloride), cetethyldimonium halides, cetylpyridinium halides, benzododecinium halides, chlorallyl methenamine halides, myristalkonium halides, stearalkonium halides and parabens.,

[0060] In this particular embodiment, the composition may comprise at most 0.01% by weight, relative to the total weight of the composition, of a microbicidal and / or microstatic agent (non-surfactant), notably selected from the group consisting of antioxidant agents, boric acid and its salts, benzoic derivatives, a salt thereof, or a mixture thereof.

[0061] According to another particular embodiment, the composition is free of preservatives, surfactant or non-surfactant, that is to say that it comprises at most 0.001% by weight of preservative, and advantageously at most 0.0001% by weight of preservatives, relative to the total weight of the composition.

[0062] Non-ionic oil-in-water emulsion

[0063] The composition comprises an oil-in-water emulsion, also referred to as O / W. Thus, the aqueous phase Aq forms the continuous phase of the O / W emulsion, while the lipid phase L (also called the oily phase) forms the discontinuous phase of the O / W emulsion. More specifically, the lipid phase is in the form of droplets dispersed in the aqueous phase. Advantageously, the lipid phase is present at a content of between 0.1% and 5%, in particular between 0.5 and 2%, for example 1%, by volume relative to the total volume of the composition.

[0064] Lipid Phase L

[0065] The lipid phase preferably comprises, or consists essentially of, glycerol esters of the following formula (I):

[0066] [Chem. 1] in which R 1 , R 2 and R 3 independently represent H, a -(O=C)-C1-C18 alkyl group or a -(O=C)-C2-C18 alkenyl group, it being understood that at least one of R 1 , R 2 and R is different from H.

[0067] For the purposes of the present invention, the term “(C1-C18) alkyl” group means a saturated, linear or branched monovalent hydrocarbon chain containing 1 to 18 carbon atoms. Examples that may be mentioned include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl or hexyl groups.

[0068] For the purposes of the present invention, the term “(C2-C18) alkenyl” group means a monovalent, linear or branched hydrocarbon chain comprising at least one double bond and comprising 2 to 18 carbon atoms. Examples of linoleyl groups include.

[0069] Preferably, R1, R2 and R3, independently of each other, represent H, a -(O=C)-C6-C8 alkyl group or a -(O=C)-C6-C8 alkenyl group, it being understood that at least one of R1, R2 and R is different from H.

[0070] According to a particular embodiment, one of R1, R2 and R3 represents a linoleyl group, and the other two residues among R1, R2 and Rs represent H. Advantageously, it is glycerol monolinoleate.

[0071] According to another particular embodiment, the lipid phase comprises or is essentially composed of triglycerides. According to this embodiment, R1, R2 and R3, independently of one another, represent a -(O=C)-C1-C18 alkyl group or a -(O=C)-C2-C18 alkenyl group. Advantageously, R1, R2 and R3, independently of one another, represent a -(O=C)-C6-C8 alkyl group.

[0072] According to a particular embodiment, the lipid phase comprises or is essentially composed of triglycerides and / or glycerol monolinoleate.

[0073] The oil phase may also include agents that affect the penetration of the active ingredient. Penetration agents that can be used are, for example:

[0074] - certain heterocyclic amine amide compounds,

[0075] - glycols, such as propylene glycol;

[0076] - propylene carbonates; or

[0077] As stated above, the emulsion is a Pickering emulsion: it is stabilized by a particulate surfactant - i.e. comprising organic particles capable of modifying the surface tension of the emulsion - and not by non-particulate surfactants.

[0078] Advantageously, the particulate surfactant / lipid phase mass ratio is between 0.1% and 2%. A person skilled in the art will be able to adapt this ratio according to the size and number of droplets desired to stabilize the emulsion while maintaining good absorption of the active ingredient where appropriate.

[0079] Preferably, the O / W emulsion is free of non-particulate surfactants, whether ionic or non-ionic. Alternatively, the emulsion is free of benzalkonium chloride, macrogolyglycerol hydroxystearate and PS80 (also known as Tween 80). The emulsion is characterized by its volume average diameter and the Span value.

[0080] The volume average diameter - noted D[4,3] - is given by equation (1):

[0081] Eq. (1): D [4 ,3] =(Z nidi 4 ) / (Z nidi 3 ) in which ni represents the number of particles denoted i with diameters dj.

[0082] The polydispersity of the droplet size distribution is expressed by the value of the “Span”, defined by equation (2):

[0083] Eq. (2): Span = (D90-D10) / D5o

[0084] Where Dw, Dso and D90 represent the diameter of the droplets below which 10%, 50% and 90% of the sample are located, respectively.

[0085] The values ​​of D[4.3], D10, D50 and D90 can be measured by LASER diffraction.

[0086] Thus, the droplets typically have a volume average diameter of less than or equal to 100 pm, preferably between 10 and 80 pm, in particular between 20 and 70 pm. Advantageously, the droplets have an average diameter of 60 pm. Indeed, droplets with excessively high average diameters can slow down diffusion, and therefore the release of the liposoluble active ingredient.

[0087] Furthermore, the droplets advantageously have a Span value between 1 and 3, particularly between 1.5 and 2. Span values ​​less than or equal to 3 are preferred, because too great a dispersion can harm the bioavailability of the active ingredient. Furthermore, emulsions with large dispersions are generally difficult to obtain in a reproducible manner.

[0088] Aqueous phase Aq

[0089] The compositions may also contain conventional, pharmaceutically acceptable additives, as listed below.

[0090] Optionally, the ophthalmic composition may contain other ingredients such as, but not limited to, osmolarity-affecting agents, buffers, or viscosity-increasing agents.

[0091] Various compounds that influence osmolarity can be used to adjust the osmolarity of a solution to approximate the composition of natural tears. For example, sodium chloride, potassium chloride, magnesium chloride, calcium chloride, dextrose, and / or mannitol can be used to approximate physiological osmolarity (generally in the range of 150-450 mOsm and preferably 250-350 mOsm).

[0092] Viscosity-acting agents (also called viscosifying agents) that may be used in the ophthalmic formulations of the present invention may be, but are not limited to, xanthan gum, gellan gum, monomeric polyols, povidone, hydroxypropylmethyl cellulose, poloxamers, carboxymethyl cellulose, carbomers or hydroxyethylcellulose, dextrans such as dextran 70, water-soluble proteins such as gelatin, hyaluronic acid or sodium hyaluronate. Preferably, it is hyaluronic acid or sodium hyaluronate.

[0093] The pH of the aqueous phase is typically between 5.5 and 7.5, for example between 5.8 and 6.5, in particular equal to 6.

[0094] According to a first variant, the aqueous phase Aq is essentially composed of water and buffers. This may in particular be water for injections (WFI water), mixed with a buffer, and optionally an osmolarity modifying agent and / or a viscosifying agent, in particular as indicated previously.

[0095] The buffer typically includes acids and / or bases. For example, sodium hydroxide (NaOH), sodium citrate, and / or acetic acid can be used as a buffer.

[0096] According to a second variant, the aqueous phase is preferably in the form of an aqueous gel.

[0097] The aqueous gel is advantageously obtained by gelling an aqueous solution comprising pharmaceutically acceptable viscosifying agents, such as those listed above, and preferably xanthan gum, gellan gum, hyaluronic acid and / or sodium hyaluronate, all cellulose derivatives (HPMC, HEC, EC, etc.) or xanthan gum.

[0098] Advantageously, the viscosifying agent is present in an amount sufficient to achieve a dynamic viscosity at rest typically between 3000 and 3500 mPa.s, advantageously between 3200 and 3300 mPa.s. Preferably, the viscosifying agent is also present in an amount sufficient to achieve a loss factor tan(ô) at rest of less than 1, preferably between 0.8 and 0.15. Those skilled in the art know how to determine the gelling agent content to achieve the targeted dynamic viscosity and loss factor tan(ô) ranges, a content which depends on the nature but also on the molecular weight of the gelling agent.

[0099] The emulsion rest loss factor tan(δ) is typically measured at 20°C by performing an oscillation amplitude sweep using a MCR102 rotary rheometer (Anton Paar GmbH, Graz, Austria) equipped with a cone-plate geometry of 50 mm diameter and 1° angle. The shear strain is varied from 0.4 to 100%. The rest loss factor tan(δ) corresponds to the value obtained in the viscoelastic linearity range.

[0100] The dynamic viscosity at rest is measured after dilution in artificial tears adjusted to pH 7.4 at a ratio of 30:7 (EmulsiomArtificial Tears), in order to simulate the dilution during administration in the eye. The flow curves are obtained using a rotational rheometer, such as an MCR102 rheometer (Anton Paar GmbH, Graz, Austria) with a cone-plate geometry of diameter 50 mm and angle 1 ° and a Peltier plane to control the sample temperature at 20 °C. The shear rate is varied from 0.1 to 1000 s-1 with an acquisition time of 10 to 1 s per point. The value retained is the initial dynamic viscosity of the flow curve. Artificial tears typically have the following compositions:

[0101] [TABLE 1]

[0102] In this second variant, the aqueous gel further increases the stabilization of the emulsion by preventing or slowing down creaming.

[0103] Creaming is a phenomenon of gravitational phase separation due to the difference in gravity of the phases: the dispersed lipid phase, less dense than the aqueous phase, migrates upwards, generating a vertical concentration gradient. Creaming is a reversible phenomenon, however, it is preferable to avoid it because it promotes flocculation, a prerequisite for the coalescence of the droplets, which increases their size irreversibly.

[0104] According to this second variant, the aqueous gel typically comprises a buffer, typically comprising acids and / or bases. For example, sodium hydroxide (NaOH), sodium citrate and / or acetic acid may be used as a buffer.

[0105] Particulate surfactant

[0106] The particulate surfactant comprises or consists of organic polymer particles.

[0107] Particles are defined by their intensity-averaged size and their polydispersity index. "Intensity-averaged size" refers to the harmonic mean diameter, and is denoted by Z-av. The intensity-averaged size can be expressed as the harmonic mean based on the intensity (2,3) and is illustrated by the equation below:

[0108] [Math 1] where Si is the scattered intensity of particle i and Dj is the diameter of particle i. The value of the average size Z is calculated by cumulant methods (see in particular J. Chem. Phys 57 (11), pp 4814-4820, 1972.). It is based on measurements carried out by dynamic light scattering (DLS).

[0109] Polydispersity index is a measure of the heterogeneity of a sample as a function of its size. Polydispersity can be due to the size distribution in a sample. Thus, in the case of a peak, the polydispersity index is 1, in the case of a Gaussian, the index can be greater than 3. The polydispersity index is generally measured by DLS.

[0110] The organic particles typically have an average size in intensity (Z-Av) less than or equal to 500 nm, in particular between 100 and 300 nm, preferably between 150 and 250 nm. Particles of 500 nm or less are preferred, because they are distributed on the surface of the droplets and confer sufficient stability to the emulsion.

[0111] The organic particles typically have a polydispersity index less than or equal to 0.3, preferably between 0.05 and 0.2, in particular between 0.1 and 0.15.

[0112] According to an advantageous variant, the particulate surfactant does not act as a vector for an active ingredient: thus, the solid particles of the particulate surfactant are devoid of active ingredient.

[0113] Advantageously, the surfactant comprises or consists of particles of a mucoadhesive polymer, preferably chosen from hyaluronic acid, chitosan, PLA-PEG ((polylactic acid)-polyethylene glycol), PLGA (poly(lactic-co-glycolic acid)), PLGA-PEG (poly(lactic-co-glycolic acid)-polyethylene glycol), PVP (Polyvinylpyrrolidone), PVP-PEG (Polyvinylpyrrolidone-polyethylene glycol), PCL (Polycaprolactone), PCL-PEG (Polycaprolactone-polyethylene glycol), and mixtures thereof.

[0114] Advantageously, these are chitosan particles. Indeed, such particles have mucoadhesive properties, beneficial for improving the residence time of the composition on the ocular mucosa, and therefore the bioavailability of the active ingredient.

[0115] The chitosan particles are obtained by coprecipitation with an organic acid salt. For the purposes of the present invention, an “organic acid” means an organic molecule comprising at least one carboxylic acid function -COOH, and whose pKa is between 3 and 5. It may in particular be an acid of formula R-COOH, in which R represents a C1-C6 alkyl (preferably C1-C4) optionally substituted by one to three substituents chosen from -OH, -COOH. Examples of organic acids according to the invention are acetic acid, lactic acid, citric acid. The organic acid is pharmaceutically acceptable, and suitable for ophthalmic applications. Alternatively, it may be tripolyphosphate (CAS No. 14127-68-5).For the purposes of the present invention, the term "organic acid salt" means the pharmaceutically acceptable base addition salts formed when an acid proton present in the organic acid is either replaced by a metal ion, for example an alkali metal ion, an alkaline earth metal ion or an aluminum ion; or coordinated with a pharmaceutically acceptable organic or inorganic base. Acceptable organic bases include diethanolamine, ethanolamine, N-methylglucamine, triethanolamine, tromethamine and the like. Acceptable inorganic bases include aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate and sodium hydroxide. Preferably, it will be a sodium salt.

[0116] Preferably, the organic acid salt is sodium hyaluronate or sodium citrate. In a particular embodiment of the invention, the surfactant comprises or consists of particles obtained by co-precipitation between chitosan and sodium citrate. Regarding the protocol for forming the particles of the surfactant, the chitosan is dissolved in water, then the solution is optionally filtered. The acid salt is added (with stirring) either in solid form or in solution form, then the pH of the solution is raised to approximately 6 by adding a base, preferably mineral and if appropriate with a cation identical to that of the weak acid salt, such as sodium hydroxide. The formation of nanoparticles is then observed.

[0117] Active ingredient

[0118] According to one embodiment, the composition comprises at least one active ingredient.

[0119] The active ingredient can be fat-soluble or water-soluble.

[0120] For the purposes of the invention, "water-soluble active ingredient" means an active ingredient having high solubility in water, within the meaning of the biopharmaceutical classification system "BCS" (cited above). More specifically, reference is made to active ingredients of class I or III within the meaning of the BCS. Thus, an active ingredient is water-soluble when the entire highest therapeutic dose of the active ingredient in question is soluble in 250 mL of a buffered aqueous solution over a pH range of 1 to 6.8 at 37°C±1°C.

[0121] According to one embodiment, the ophthalmic compositions of the invention contain an effective amount of water-soluble active ingredient.

[0122] Examples of water-soluble active ingredients are beta-blockers, in particular chosen from timolol, levobunolol, dorzolamide and / or brinzolamide, preferably timolol.

[0123] In this embodiment, the water-soluble active ingredient is advantageously included in the aqueous phase Aq, or is adsorbed on the surface of the organic particles. However, the active ingredient does not form a covalent bond with the organic particles. The formulation of Pickering emulsions with a water-soluble active ingredient is well known to those skilled in the art. According to one embodiment of the invention, the composition, and more particularly the lipid phase L, comprises at least one liposoluble active ingredient.

[0124] “Fat-soluble active ingredient” means an active ingredient with low solubility in water, within the meaning of the biopharmaceutical classification system “BCS”, as defined in particular in the document “M9 Biopharmaceutics Classification SystemBased Biowaivers - Guidance for Industry” published in May 2019 by the American “Food and Drug Administration” (FDA), or the document “ich-m9-biopharmaceutics-classification-system-based-biowaivers-step-5” officially published by the European Medicines Agency (EMA) on 07 / 30 / 2020, and available online at: https: / / www.ema.europa.eu / documents / scientific-guideline / ich-m9-biopharmaceutics-classification-system-based-biowaivers-step-5_en.pdf. More specifically, we are referring to active ingredients of class II or IV within the meaning of the BCS.Thus, an active ingredient is considered fat-soluble when less than 85% by weight of the highest therapeutic dose is soluble in 250 mL of a buffered aqueous solution over a pH range of 1 to 6.8 at 37°C±1°C.

[0125] The ophthalmic compositions contain an effective amount of fat-soluble active ingredient. Generally, the compositions described herein will contain between 5 pg / mL and 10 mg / mL of fat-soluble active ingredient.

[0126] Examples of fat-soluble active ingredients include prostaglandins and steroids.

[0127] By "prostaglandins" is meant compounds of the prostaglandin family, their derivatives and / or analogues, whether of natural or synthetic origin, such as prostamides. Prostaglandins have a cyclopentyl substituted in positions 1 and 4 by oxygen atoms (e.g. by two hydroxyl groups, or carrying a C=O ketone function in position 1 and substituted by a hydroxyl group in position 4), and in positions 2 and 3 by two identical or different monovalent hydrocarbon chains, saturated or unsaturated (i.e. comprising at least one C=C double bond), linear or branched, comprising 1 to 12 carbon atoms. The chain in position 3 generally comprises an ester or amide function. Prostaglandins can be of series H, D, E, F, Alpha or I and of class 1, 2 or 3.

[0128] Advantageously, prostaglandins with therapeutic indications for the treatment or prevention of ophthalmic diseases or conditions will be chosen. In particular, prostaglandins are selected from the group consisting of latanoprost, travoprost, bimatoprost and tafluprost.

[0129] When the liposoluble active ingredient is a prostaglandin, the compositions of the invention will typically contain between 5 pg / mL and 500 pg / mL, in particular from 20 to 400 pg / mL, for example from 40 to 300 pg / mL, of liposoluble active ingredient. Preferably, for Latanoprost or Travoprost, the composition comprises between 10 and 100 pg / mL, in particular between 20 and 75 pg / mL, for example 50 pg / ml or 40 pg / ml of liposoluble active ingredient.

[0130] Preferably, for Latanoprost, the composition comprises between 10 and 100 pg / mL, in particular between 20 and 75 pg / mL, for example 50 pg / ml of liposoluble active ingredient.

[0131] Advantageously, for Bimatoprost, the composition comprises between 10 and 100 pg / mL, in particular between 50 and 400 pg / mL, for example between 100 and 300 pg / ml of liposoluble active ingredient.

[0132] "Steroids" means a compound comprising or deriving from a partially or fully hydrogenated hydrophobic cyclopentanophenanthrenic (sterane) nucleus. Steroids include, in particular, sterols characterized by the presence of a hydroxyl group on the C carbon 3 (e.g., cholesterol).

[0133] Steroids are or behave like hormones. They can be grouped into five categories based on their receptors: glucocorticoids, mineralocorticoids, androgens, estrogens, and progestins. Vitamin D derivatives form a sixth hormonal system.

[0134] • Glucocorticoids: cortisol, dexamethazone

[0135] • Mineralocorticoids: aldosterone, fludrocortisone

[0136] • Androgens: testosterone, dehydroepiandrosterone (DHEA), dehydroepiandrosterone sulfate (DHEAS), androstenedione, dihydrotestosterone (DHT)

[0137] • Estrogens: estradiol, estrone, estriol

[0138] • Progestins: progesterone

[0139] Advantageously, a steroid chosen from steroidal anti-inflammatory drugs, such as hydrocortisone, prednisone, prednisolone, methylprednisolone, betamethasone, dexamethasone, triamcinolone, estradiol, estrone, estriol, progesterone, and mixtures thereof, will be chosen.

[0140] When the liposoluble active ingredient is a corticosteroid, the compositions of the invention will typically contain between 0.01 mg / mL and 10 mg / mL, in particular from 0.05 to 7.5 mg / mL, for example from 0.1 to 5 mL, of liposoluble active ingredient.

[0141] Advantageously, for dexamethasone, the composition comprises between 0.1 and 2 mg / mL, for example 1 mg / ml of liposoluble active ingredient.

[0142] In a particular embodiment, the composition comprises at least one liposoluble active ingredient and optionally a water-soluble active ingredient. For example, in a particular embodiment of the invention, the composition comprises both a liposoluble active ingredient, such as a prostaglandin (PGA) derivative or a steroid derivative, and a water-soluble active ingredient such as a β-blocking agent. According to a particular embodiment, the composition comprises one or more protaglandin(s) in combination with a beta-blocker chosen from timolol, levobunolol, dorzolamide and / or brinzolamide, preferably timolol. In this embodiment, the prostaglandin is advantageously included in the lipid phase, while the β-blocker is advantageously included in the aqueous phase.

[0143] Special embodiments

[0144] According to one embodiment, the ophthalmic composition comprises a non-ionic oil-in-water emulsion with:

[0145] • an aqueous phase Aq,

[0146] • a lipid phase L, in the form of droplets dispersed in the aqueous phase,

[0147] • an interface between said lipid phase L and said aqueous phase Aq, and

[0148] • a surfactant for stabilizing the emulsion, located at the interface between the lipid phase and the aqueous phase, said lipid phase L comprising at least one liposoluble active ingredient, said surfactant being particulate, in the form of organic particles of average size in intensity between 100 and 300 nm, preferably between 150 and 250 nm, and with a polydispersity index typically less than or equal to 0.3, preferably between 0.05 and 0.2, in particular between 0.1 and 0.15.

[0149] According to this embodiment, the lipid phase is present at a content of between 0.1% and 5%, in particular between 0.5 and 2%, for example 1%, by volume relative to the total volume of the composition. The droplets of the emulsion advantageously have a volume average diameter of less than or equal to 100 μm, preferably between 10 and approximately 80 μm, in particular between 20 and 70 μm, and a Span value of between 1 and 3, in particular between 1.5 and 2.

[0150] Typically, the lipid phase preferably comprises, or consists essentially of, glycerol esters of formula (I): [Chem. 1] in which R 1 , R 2 and R 3 independently represent H, a -(O=C)-C1-C18 alkyl group or a -(O=C)-C2-C18 alkenyl group, it being understood that at least one of R 1 , R 2 and R is other than H, and may also comprise penetrating agents.

[0151] Preferably, the surfactant comprises or consists of chitosan particles. Advantageously, the particulate surfactant / lipid phase mass ratio is between 0.1% and 2%.

[0152] Typically, the pH of the aqueous phase is typically between 5.5 and 7.5, for example between 5.8 and 6.5, and the aqueous phase is preferably in the form of an aqueous gel.

[0153] The aqueous gel is for example obtained by gelling an aqueous solution comprising pharmaceutically acceptable viscosifying agents, such as xanthan gum, gellan gum, hyaluronic acid and / or sodium hyaluronate, cellulose derivatives, xanthan gum and alginate. The viscosifying agent is present in an amount sufficient to achieve a dynamic viscosity at rest typically between 3000 and 3500 mPa.s, advantageously between 3200 and 3300 mPa.s. Preferably, the viscosifying agent is also present in an amount sufficient to achieve a loss factor tan(5) at rest of less than 1, preferably between 0.8 and 0.15.

[0154] Preferably, the active ingredient is chosen from prostaglandins (such as latanoprost, travoprost, bimatoprost and tafluprost), optionally in admixture with timolodol, levobunolol, dorzolamide and / or brinzolamide, and steroids (such as hydrocortisone, prednisone, prednisolone, methylprednisolone, betamethasone, dexamethasone, triamcinolone, and mixtures thereof), at a concentration of between 5 pg / mL and 10 mg / mL of prostaglandins.

[0155] Process for preparing the emulsion

[0156] The composition is prepared according to methods known to those skilled in the art.

[0157] The composition of the invention, which comprises:

[0158] • an aqueous phase Aq,

[0159] • a lipid phase L, in the form of droplets dispersed in the aqueous phase, comprising at least one liposoluble active ingredient,

[0160] • an interface between said lipid phase L and said aqueous phase Aq, and

[0161] • a particulate surfactant, in the form of organic particles of average size in intensity less than or equal to 500 nm to stabilize the emulsion, located at the interface between the lipid phase and the aqueous phase, is advantageously prepared according to the following steps: a) Suspending the particulate surfactant in the aqueous phase, b) Dissolving the active ingredient in an oil to form the lipid phase, c) Adding the lipid phase to the suspension obtained in step a), d) Emulsification, so as to obtain an emulsion. The droplets of the emulsion obtained at the end of step d) typically have a volume average diameter less than or equal to 100 μm, preferably between 10 and approximately 80 μm, in particular between 20 and 70 μm.

[0162] Furthermore, the droplets advantageously have a Span value between 1 and 3, in particular between 1.5 and 2.

[0163] Preferably, the emulsification step d) is carried out using a high shear mixer, such as a rotor / stator mixer, or a high pressure homogenizer. The emulsification is preferably carried out without heating, and / or at room temperature. When the mixer is a rotor / stator, it typically operates at a stirring speed greater than 10,000 rpm, for example between 10,000 and 20,000 rpm.

[0164] During step c) of adding the lipid phase to the aqueous phase, the lipid phase preferably represents between 0.1% and 5%, in particular between 0.5 and 2%, for example 1%, by volume relative to the total volume of the composition. Step c) can be carried out in a conventional manner for those skilled in the art. Step c) is typically carried out at room temperature.

[0165] The oil used in step b) of dissolving the active ingredient in an oil to form the lipid phase is as previously described in the lipid phase L section, and comprises, or consists essentially of, glycerol esters of formula (I) as defined above, and optionally penetrating agents. Step b) is typically carried out at room temperature.

[0166] The aqueous phase comprises or consists essentially of water (in particular WFI water), and optionally of agents influencing osmolarity, buffers, and / or viscosifying agents, as defined above.

[0167] The aqueous phase used in step a) is preferably gelled. In this case, the method comprises a prior step of gelling an aqueous solution, said gelling step comprising: a1) dissolving a gelling agent in water, typically with stirring and preferably heating to a temperature between 50 and 80°C then cooling to room temperature, a2) optionally adding agents influencing osmolarity, buffers, and / or viscosifying agents in solution in water, a3) homogenizing the gelled aqueous solution to form the aqueous phase.

[0168] The quantities of water gelling agent, and where appropriate other optional additives, are chosen so as to achieve a dynamic viscosity at rest typically between 3000 and 3500 mPa.s, advantageously between 3200 and 3300 mPa.s and preferably a loss factor tan(5) at rest of less than 1, preferably between 0.8 and 0.15. Therapeutic Applications

[0169] The ophthalmic compositions of the invention are useful as medicaments, particularly for administration by ocular instillation.

[0170] According to a first variant, the composition is useful for the treatment of dry eye. In this case, the composition does not necessarily contain an active ingredient, the oily phase and, where appropriate, the gelling agent (in particular sodium hyaluronate) being sufficient in itself to combat dry eye.

[0171] According to a second variant, the composition comprises a liposoluble active ingredient.

[0172] According to one embodiment, the active ingredient is selected from steroids. These particular compositions are useful as medicaments, in particular for preventing, relieving or treating ocular inflammations. Alternatively, the invention relates to a method for preventing, relieving or treating ocular inflammations, comprising the ocular administration of an effective amount of the ophthalmic composition to a patient in need thereof, the active ingredient being selected from steroids.

[0173] Ocular inflammation can cause, among other things, a breakdown of the barriers that isolate the eye from the general circulation, allowing the entry of proteins and cells that participate in the inflammatory process and have an impact on healing. Inflammation can affect the cornea and sclera on the surface of the eye; the uvea composed of the iris, ciliary body and choroid; and the origin of the optic nerve, the retina and the papilla. For example, ocular inflammation affects the sclera on the surface of the eye; the uvea composed of the iris, ciliary body and choroid; and the origin of the optic nerve, the retina and the papilla.This ocular inflammation can be responsible for opacities and lesions of the cornea (keratitis); lesions of the sclera (scleritis and episcleritis); uveitis with the complication of opacification of the lens (cataract); adhesions of the iris to the lens (synechiae); macular edema; retinal detachment; ocular hypertension and degeneration of the optic nerve caused by glaucoma; the formation of new vessels (neovascularization); etc.

[0174] According to another embodiment, the active ingredient is chosen from prostaglandins, optionally in admixture with timolodol, levobunolol, dorzolamide and / or brinzolamide. These particular compositions are useful as medicaments, in particular in the treatment or prevention of intraocular hypertension, and / or glaucoma, such as open-angle glaucoma. Alternatively, the invention relates to a method for preventing or treating intraocular hypertension, and / or glaucoma, such as open-angle glaucoma, comprising the ocular administration of an effective amount of the ophthalmic composition, the active ingredient being chosen from prostaglandins, optionally in admixture with timolodol, levobunolol, dorzolamide and / or brinzolamide.

[0175] For the purposes of the present invention, an "effective amount" refers to the level or quantity of active ingredient targeted, without causing significant negative or undesirable side effects on the target, (1) delaying or preventing the onset of the disease or condition; (2) slowing or stopping the progression, aggravation, or deterioration of one or more symptoms of the disease or condition; (3) causing improvements in the symptoms of the disease or condition; (4) reducing the severity or incidence of the disease or condition; or (5) curing the disease or condition. An effective amount may be administered before the onset of the disease or condition, for prophylactic or preventive action. Alternatively or in addition, the effective amount may be administered after the initiation of the disease or condition, for therapeutic action.

[0176] The patient can be a mammal, preferably a human.

[0177] FIGURES

[0178] [Fig.1] Figure 1 schematically illustrates the structure of the three emulsions NP-E, GE and NP-GE of Example 3. The spheres represent the droplets of the lipid phase, while the network represents the gel form of the gelled aqueous phase.

[0179] [Fig.2] Rheological characterization of the emulsions of interest. A) Flow curves representing the dynamic viscosity as a function of the shear rate. B) Plot of the shear stress as a function of the shear rate. C) Blink simulation test over 10 cycles of 1 min. D) Visual observation of creaming after 8 weeks.

[0180] [Fig.3] Particle size measurements of emulsions obtained by laser diffraction. A) Volume distributions for the three emulsions studied, NP-E, GE, and NP-GE at the initial time. B) Evolution of D[4,3] over time after storage at room temperature. The results are represented by the mean and standard deviation of the measurements of 3 independent samples (n = 3).

[0181] [Fig.4] Travoprost release profiles for Travatan® (- -), NP-E simple Pickering emulsion (-), GE emulsion / gel (-) and Pickering emulsion in NP-GE gel (-).

[0182] [Fig.5] Relative cell viability of the three emulsions selected in comparison with commercial Travatan®. BAK: Benzalkonium Chloride. **p-value < 0.01 in a t-test with Welch correction.

[0183] EXAMPLES

[0184] ABBREVIATIONS

[0185] ACN acetonitrile rpm rotation per minute

[0186] Tamb Room temperature

[0187] PPI For Injectable Preparations v / v volume / volume m / v mass / volume

[0188] DLS analysis by LASER diffraction (in English “Dynamic Light Scattering”)

[0189] Z-Avg Average size in intensity

[0190] PDI Polydispersity Index

[0191] PS80 PolySorbate 80

[0192] NaHA Sodium Hyaluronate

[0193] Cs Chitosan

[0194] Qsp Quantity sufficient for

[0195] MATERIALS AND METHODS

[0196] Three types of Cs (Chitoclear®) with 1% w / v solution viscosities of 10, 12, and 90 mPa.s (supplier's information) and deacetylation degrees of 95, 74, and 95% (supplier's information) respectively were supplied by Primex (Siglufjôrôur, Iceland). Glacial acetic acid grade AnalaR® NORMAPUR® was purchased from VWR Chemicals (Radnor, PA, USA). 2N sodium hydroxide was supplied by LCH Chimie (Les Aires, France). Pharmaceutical excipient grade sodium hyaluronate (NaHA) with an intrinsic viscosity of 2.5 m3 / kg was supplied by Shiseido CO., LTD. (Kakegawa-shi, Japan). Sodium citrate dihydrate (molar mass 294.1 g / mol) was supplied by Caesar & Loretz GmbH (Hilden, Germany). Gellan gum (Kelcogel® CG-LA) was supplied by CP Kelco (Atlanta, GA, USA). Lipophilic Labrafac™ WL 1349 (Medium Chain Triglycerides, MCT) was supplied by Gattefossé (Lyon, France).Travoprost was provided by Sanofi Chinoin (Budapest, Hungary). Pyrene of 98% purity was provided by Acros Organics (lllkirch, France). Rhodamine-grafted Cs (Cs-Rhodamine), with a molecular weight of 5 kDa and a degree of deacetylation of 95%, and a molar substitution of 1%, was provided by HAworks LLC (Bridgewater, NJ, USA). Sodium chloride (NaCl), calcium chloride dihydrate (CaCl2 2H2O), magnesium chloride (MgCl2), potassium chloride (KCl), sodium bicarbonate (NaHCO3), ACN, and porcine mucin type II were purchased from Sigma-Aldrich (St. Louis, MO, USA). The commercial product Travatan® was provided by Alcon® (Hünenberg, Switzerland). Lavoisier water for injection (WFI) was supplied by CDM Lavoisier Laboratories (Paris, France).

[0197] Granulometric analyses by laser diffraction

[0198] The colloidal stability of the emulsions over time after storage at room temperature was evaluated by laser diffraction. After morphological observation under a transmission optical microscope, the D[4.3] of the globules was measured by laser diffraction using a MasterSizer 3000 equipped with a Hydro SM liquid cell (Malvern Panalytical Ltd., Malvern, UK). The measurements were carried out under mechanical stirring at 2500 rpm at 10 and 15% obscuration. For the processing of the acquired data, the particle type chosen was spherical, with a refractive index of 1.47 and an absorption index of 0.002 as previously determined by the optical properties optimizer of the Mastersizer software Version 3.62 (Malvern Panalytical Ltd., Malvern, UK).The result for a sample consists of the average of 6 measurements of 10 s spaced 30 s apart, ensuring compliance with the variability criteria of the European Pharmacopoeia (< 10% for D50 and < 15% for D10 and D90).

[0199] Observation by fluorescence microscopy

[0200] The probes used are pyrene for the lipid phase, and Cs-rhodamine of similar molecular weight and degree of deacetylation for the solid nanoparticles.

[0201] For the purpose of measurements, samples are formed as follows: pyrene is dissolved in the lipid phase to obtain a concentration of 4 mg / mL prior to emulsification, and 1% of the mass of Cs used to formulate the NPs is substituted by Cs-rhodamine of similar molecular weight and degree of deacetylation.

[0202] Transmission and fluorescence observations were performed using an Axiophot fluorescence microscope (Zeiss, Oberkochen, Germany) equipped with an X-Cite® 120Q light source (Excelitas Technologies, Waltham, MA, USA). Image acquisition was performed using a Qlmaging® Retiga-2000R digital camera (Teledyne Photometries, Tucson, AZ, USA) and QCapture Pro software version 6.0.0.605 (Teledyne Photometries, Tucson, AZ, USA). The filters used to view pyrene and Cs-rhodamine were exc. BP 365 / 12 em. LP 397 and exc. 546 / 12 em. LP 590, respectively.

[0203] The images obtained were analyzed using ImageJ version 1.53s software from the National Institutes of Health (Madison, WI, USA).

[0204] USP4 release studies

[0205] The release of travoprost from the compositions was evaluated using a USP4 CE7 Smart dissolution apparatus coupled with a CP7 pump (Sotax, Aesch, Switzerland). 22.6 mm diameter cells were used. A 5 mm ruby ​​bead was placed at the bottom of the cells, then covered by 1 mm diameter glass beads. This arrangement ensured a laminar flow of the dissolution medium at the cell level. Artificial tears adjusted to a pH of 7.4 were used as the dissolution medium, with the addition of a 0.1% v / v concentration of PS80 to satisfy closed-circuit sink conditions. The volume of dissolution medium was set at 50 mL, with a pulsatile flow rate of 16 mL / min. The temperature of the dissolution medium was regulated at +35 ± 0.5 °C by the CE7 Smart apparatus water bath to mimic the temperature conditions of the ocular surface.

[0206] The emulsion test sample is 2 mL (equivalent to a dose of 80 pg of travoprost). At each time point, 1 mL of the dissolution medium was taken before adding 1 mL of new medium. The samples are immediately stored in the freezer (20 °C) for one week before performing the travoprost assay using the HPLC-UV method described in 1.7.3 below. Gompertz modeling is applied to the dissolution profiles of the emulsions from the initial time up to 85% release using DDsolver software.

[0207] Travoprost assay by HPLC-UV

[0208] Sample preparation before assay consists of a half-fold dilution in ACN, followed by centrifugation for 20 minutes at 10,000 rpm. The supernatant is collected, and 10 μL are injected for analysis. Chromatographic separation is performed using an UltiMate 3000 HPLC system (Thermo Fisher Scientific, Waltham, MA, USA) equipped with a Kinetex C18 bonded silica column with dimensions of 150 x 4.6 mm and particle size of 5 μm (Phenomenex, Torrance, CA, USA) set at 40 °C. The mobile phase flow rate is set at 1 mL / min. The ACN / 10 mM phosphate buffer pH 3 content of the mobile phase is varied according to the following gradient:

[0209] • from 0 to 26 minutes of analysis: linear gradient of ACN / Phosphate buffer ranging from 38 / 62 v / v to 50 / 50 v / v,

[0210] • then, from 26 to 29 minutes of analysis: isocratic phase at an ACN / Phosphate buffer ratio of 50 / 50 v / v,

[0211] • finally, 29 to 40 minutes of analysis: linear gradient of ACN / Phosphate buffer ranging from 50 / 50 v / v to 38 / 62 v / v.

[0212] Travoprost was detected using a UV detector at a wavelength of 200 nm. The chromatograms obtained were analyzed using Chromeleon™ 7.3.1 software (Thermo Fisher Scientific, Waltham, MA, USA). The calibration curve ranged from 5 to 25 pg / mL for the determination of eye drops (finished product) and from 0.082 to 2.5 pg / mL for the determination in dissolution samples. Linearity was confirmed by a coefficient of determination R2 > 0.995 and a recalculated bias for each point of the standard line of less than 5%. The accuracy of the method is verified by its precision, located between 98 and 102% as estimated from the recovery of control samples at the target concentration (20 .ig / mL) and its precision, confirmed by a coefficient of variation lower than 3% for the control samples. Finally, the repeatability of the injection was lower than 0.4%.

[0213] Cell viability assessment

[0214] The evaluation was performed on human corneal epithelial cells (immortalized HCE-T cell line). The cells were cultured under standard conditions (humid atmosphere, 5% CO2 at 37 °C) in Dulbecco's Minimum Essential Medium (DMEM) supplemented with 2.5% fetal bovine serum (FBS), 2 mM L-glutamine, 50 IU / mL penicillin, and 50 IU / mL streptomycin. The culture medium was changed every 3 days. When the cells reached confluence, they were trypsinized and then reintroduced into 96-well plates (200 pL per well) at a density of 110,000 cells / mL. Cultures are kept for 24 hours at 37°C before exposure to the compositions to allow the cells to adhere to the microplate and reach confluency.

[0215] Before incubation, the cells were washed with 200 pL of phosphate buffer saline (Phosphate Buffer Saline, PBS), then the compositions were incubated neat (100 pL / well) for 30 minutes on the cells.

[0216] After incubation, the composition is removed and the cells are washed again with 200 μL of PBS, then incubated with DMEM culture medium supplemented with 2.5% FCS for 24 hours. AlamarBlue™ dye solution (200 μL) is added to each well for incubation for 6 hours at 37°C. The AlamarBlue™ dye solution is pre-diluted to 0.09 mg / mL in the culture medium supplemented with 2.5% FCS. The fluorescence of AlamarBlue™ (λex = 535 nm; λem = 600 nm) is measured using a cytofluorometer (Safire, Tecan Group Ltd., Mannedorf, Switzerland) adapted to microplates. The fluorescence intensity results obtained are expressed as the percentage of viable cells compared to the control.

[0217] 0.01% BAK was used as a positive control. Results presented are the mean and standard deviation of 3 independent runs.

[0218] EXPERIMENTAL PART

[0219] Example 1a - Preparation of Chitosan-sodium hyaluronate (NaHA) nanoparticles Cs was dissolved in a 0.1% v / v acetic acid solution under magnetic stirring at 350 rpm for at least 12 h before being filtered on a Clearline® polyethersulfone (PES) membrane with a porosity of 0.45 pm (Dutscher, Bernolsheim, France). On the other hand, NaHA was dissolved in WFI water under magnetic stirring at 600 rpm for at least 6 h. Both solutions were left to stand overnight. 2N NaOH was added to the NaHA solution to obtain a pH of 6.0 at the end of the process.

[0220] The NaHA solution was then gradually added into the Cs solution, by a continuous flow under stirring.

[0221] The nanoparticles thus obtained have a Z-Avg of 190 nm and a PDI of 0.159.

[0222] Example 1b - Preparation of Chitosan Nanoparticles - Citrate

[0223] Chitosan with a 1% m / v solution viscosity of 10 mPa.s and a deacetylation degree of 95% and sodium citrate are used.

[0224] Chitosan was dissolved in a 0.1% v / v acetic acid solution to reach a concentration of 0.04% w / v (12 h of magnetic stirring at room temperature at 350 rpm). The resulting solution was filtered through a 0.45 μm polyethersulfone (PES) filter before neutralizing the pH to 6.0 with 0.4 N NaOH. Sodium citrate was dissolved in the filtered solution to reach a concentration of 2.94% w / v. Acetic acid was then added from a 5% v / v solution under magnetic stirring at 500 rpm until a pH of 6.0 was reached. The resulting nanoparticles had a Z-avg of 193 nm and a PDI of 0.1.

[0225] For variations in chitosan and sodium citrate concentrations between 0.025 and 0.04% w / v and 2.5 and 3.0% w / v respectively, the nanoparticles obtained have a Z-Avg that can vary from 180 to 200 nm and a PDI of 0.10 to 0.15 as determined by DLS measurements.

[0226] Compared to the nanoparticles of Example 1a, the size of the nanoparticles obtained is not significantly different but the PDI is clearly improved, which can be attributed to having replaced a high molar mass polymer (NaHA) with a small molecule (citrate).

[0227] Example 2 - Preparation of an emulsion according to the invention with Solid Chitosan Nanoparticles obtained according to Example 1

[0228] Preparation of the lipid phase

[0229] The oily phase of the emulsion corresponds to a 0.4% w / v solution of travoprost in medium-chain triglycerides (consisting of compounds of general formula (I) as defined above, with substituents R1, R2 and R3 chosen from C1-C18 alkyls). Dissolution is carried out at room temperature with magnetic stirring at 250 rpm for 2 h.

[0230] Preparation of a gelled aqueous phase

[0231] The aqueous phase of the emulsion comprises the nanoparticles obtained in Example 1 and a weak gel of gellan gum and sodium hyaluronate. The gel is obtained by first dissolving the gellan gum in WFI water in a water bath at 80°C under magnetic stirring at 700 rpm (gellan gum concentration of 0.6% w / v). Once dissolved, the solution is allowed to cool to room temperature. The sodium hyaluronate and the remaining WFI water are added to obtain a gellan gum and sodium hyaluronate concentration of 0.3 and 0.2% w / v, respectively. The solution is homogenized for 5 h at room temperature with magnetic stirring at 700 rpm.

[0232] Suspension of nanoparticles

[0233] The nanoparticle suspension of example 1a or 1b and the aqueous phase - preferably gelled - are mixed in equal volume to obtain the final aqueous phase of the emulsion (1 h at 1400 rpm).

[0234] Emulsification

[0235] The final emulsion is obtained by adding 1% v / v of oil phase into the aqueous phase under magnetic stirring at 1400 rpm to homogenize and stirring with a rotor-stator at 15,000 rpm for 3 minutes to emulsify the oil phase. The emulsification parameters can vary from 10,000 to 20,000 rpm and from 2 to 5 minutes without significant impact on the final size distribution of the emulsion obtained.

[0236] Results obtained for Pickering emulsions with Cs-NaHA nanoparticles

[0237] Several concentrations of 1 to 4% v / v of oil were explored.

[0238] We obtain a volume average diameter Ü4.3 varying from 32 pm to 96 pm and a Span value varying from 1.3 to 2.6 depending on the concentration of the lipid phase concentration.

[0239] Laser diffraction size measurements after emulsification showed that the oil concentration impacted the D[4,3] of the resulting emulsions. Indeed, for a higher concentration (4% vs 1% v / v) the D[4,3] of the emulsions was three times larger and the size distribution broader than for a lower oil concentration.

[0240] This observation indicates that Cs-NaHA NPs are not in excess compared to the oil.

[0241] Coalescence is observed for the 4% emulsion. Creaming is also observed for the 2% emulsion, as well as flocculation. Flocculation is also observed after 48 hours of standing for the emulsion containing 1% v / v of triglycerides (lipid phase).

[0242] A first attempt consisted of increasing the NP / oil ratio by decreasing the triglyceride concentration to 0.25% v / v. This appears to have reduced flocculation, which nevertheless persists.

[0243] Results obtained for Pickering emulsions with Cs-citrate nanoparticles

[0244] We obtain a volume average diameter Ü4.3 of approximately 60 pm and a Span value of approximately 1.9.

[0245] Furthermore, the organization of NPs around oil droplets was visualized by fluorescence microscopy, confirming their involvement in emulsion stabilization.

[0246] The absence of significance of the impacts of the different factors studied (time and speed of homogenization) on the values ​​of D[4,3] and Span allowed us to conclude that the process was robust.

[0247] Despite the stability of the emulsion, a creaming phenomenon was observed. Although this creaming is reversible and the flocculation problem previously encountered with the nanoparticles of Example 1a does not occur, creaming can promote coalescence over time. Thus, the rheology of the aqueous phase was optimized by gelation using gellan gum, which makes it possible to obtain weak gels in the presence of cations, introduced here with sodium citrate.

[0248] The evaluation of the different gels was carried out by combining a visual bottle test with rheology measurements to visually assess creaming by observing the accumulation of oil at the emulsion meniscus. A 0.05% w / v gellan gel seems sufficient to stabilize the emulsion against creaming without excessively increasing the dynamic viscosity of the medium. However, a syneresis phenomenon is observed. The addition of NaHA, a hygroscopic polymer, allows water to be retained in the gel and limits this phenomenon. The optimal gel is composed of 0.15% w / v gellan gum in association with 0.1% w / v NaHA.

[0249] For the rest of the work, 3 emulsions were selected and compared (Figure 1):

[0250] 1) a Pickering emulsion according to the invention, designated NP-E, not containing gelling polymers in the aqueous phase,

[0251] 2) a comparative emulsion stabilized only by a weak gel present in the aqueous phase, designated by GE,

[0252] 3) a Pickering emulsion according to the invention obtained with a gelled aqueous phase, noted NP-GE.

[0253] These three emulsions were compared on the basis of their colloidal stability, rheological properties, and in vitro dissolution profiles.

[0254] 1. Colloidal stability

[0255] The colloidal stability of the emulsions at room temperature was evaluated by laser diffraction, the results are presented in Figure 3. For the 3 emulsions obtained the particle size distribution (Figure 2A) can be considered as monodisperse (Span < 2) with respective spans of 1.6, 1.8 and 1.9 for the NP-E, GE and NP-GE emulsions.

[0256] The evolution of D[4,3] over time (Figure 3B) shows that despite some variations the size of the oil droplets is stabilized by the Cs-Citrate NPs and / or by the freezing of the aqueous phase.

[0257] 2. Rheology

[0258] A rheological characterization was carried out on the different emulsions (Figure 2). The flow curves (Figure 32A) show a shear-thinning behavior of the 3 formulas through the decrease in dynamic viscosity as a function of the shear gradient. It is possible to distinguish the simple Pickering emulsion, NP-E, from the GE and NP-GE emulsions containing the weak gel providing a higher dynamic viscosity. Although they all have a shear-thinning character, another notable difference can be highlighted by this rheological characterization. Indeed, by representing the shear stress as a function of the shear gradient (Figure 2B), it is possible to evaluate the yield threshold of the formulas, which is determined by the ordinate at the origin of the graph. The GE and NP-GE emulsions are again distinguished from the NP-E emulsion by their shear stress at the origin being 35 times higher.This is visually reflected by creaming of the oil droplets for the NP-E emulsion while this phenomenon is inhibited for emulsions presenting the gel in its aqueous phase (Figure 2D). Finally, a rheological characterization, linked to the thixotropy of the formulas, was carried out in order to evaluate their behavior once instilled in the eye. The evolution of the dynamic viscosity during 10 cycles mimicking the eyelid blink is shown in Figure 2D. For the three emulsions studied, a decrease in the dynamic viscosity is observed over time, which testifies to their shear-thinning and thixotropic character. On this criterion, it is the emulsions containing NP, NP-E and NP-GE, which are distinguished from the G-E formula by higher recovery indices (RI), of 66 and 60% against 49%.

[0259] 3. In vitro dissolution

[0260] The release profiles of travoprost obtained after the in vitro dissolution tests are shown in Figure 4, as well as the parameters of the Gompertz modeling. The profile of the NP-E emulsion (without gel) is similar to that of the commercial comparator, Travatan®. Indeed, more than 90% of the dose is released after 5 minutes without kinetic control. In addition, the release profiles of the emulsions with gel (GE and NP-GE) demonstrate kinetic control by the latter. The application of the Gompertz modeling allows us to assess two quantities describing the dissolution profile. First, parameter a, giving the proportion of undissolved travoprost in the release medium after 1 h, and parameter 13, evaluating the dissolution rate and dictating the shape of the profile. For the two parameters studied, there is no significant difference between the GE and NP-GE emulsions.Indeed, parameter a indicates that 13 and 16% of undissolved travoprost remains respectively after one hour, and parameter 13 is equivalent in both cases, at 1.4. NPs therefore do not appear to have an impact on the release profiles of travoprost.

[0261] 4. Assessment of cell viability

[0262] The results of the cell viability study after exposure to the 3 emulsions in comparison with commercial Travatan® are presented in Figure 5. The positive and negative controls reinforce the reliability of the test. Only 0.01% BAK induces a significant decrease in relative cell viability according to a t-test with Welch correction. These results seem to show an equivalence between the emulsions and travatan®, preserved with polyquaternium-1, from the point of view of cell viability after a punctual exposure to the different formulas.

[0263] 5. Conclusions

[0264] From the point of view of colloidal stability, although the NP-E emulsion exhibits creaming, laser diffraction measurements demonstrate effective emulsion stabilization by the Cs-Citrate NPs over a period of 6 months at room temperature. The GE and NP-GE emulsions are also stable over this period, with the gel also preventing creaming of the emulsion.

[0265] For all 3 emulsions, a shear-thinning behavior is observed, which is in agreement with the usual rheology of polymeric emulsions. In addition, the threshold fluid character obtained by the gellan gum and NaHA gel was preserved when adding the Cs-Citrate NPs (GE vs. NP-GE), which explains the satisfactory stabilization against creaming of the emulsion. During in vitro dissolution tests, the GE and NP-GE formulas slowed down the release kinetics of travoprost, compared to the Travatan® ophthalmic solution. However, this slowing down of the release of Travoprost® is not necessarily an advantage for the ocular route when the residence time of the eye drops on the eye is short.

[0266] A repeated blink simulation test was therefore carried out to evaluate the evolution of the dynamic viscosity of the different emulsions when they undergo strong cyclic stresses. This method made it possible to highlight that the NP-GE emulsion was more resistant to this type of stimulation, with a higher dynamic viscosity than the NP-E and GE emulsions at the end of the test.

[0267] In vitro cell viability tests showed equivalence between the three emulsions and the commercial Travatan® formula. However, this test is based on a single 30-minute exposure. This allows us to conclude for the moment that the formulas are relatively well tolerated at the time of their administration.

[0268] CONCLUSIONS

[0269] Compositions according to the invention have been prepared, with travoprost as the liposoluble active ingredient. They make it possible to dispense with the use of non-particulate surfactants, which guarantees good tolerance as demonstrated by cytotoxicity tests, without compromising the stability of the emulsions at room temperature.

Claims

CLAIMS 1. Ophthalmic composition, comprising an oil-in-water emulsion with: • an aqueous phase Aq, • a lipid phase L, in the form of droplets dispersed in the aqueous phase, • an interface between said lipid phase L and said aqueous phase Aq, and • a particulate surfactant for stabilizing the emulsion, located at the interface between the lipid phase and the aqueous phase, said surfactant comprising or consisting of organic particles of average size less than or equal to 1 μm, said composition being free of non-particulate surfactant, and advantageously free of preservative.

2. Composition according to claim 1, characterized in that the droplets have a volume average diameter less than or equal to 100 pm, preferably between 10 and approximately 80 pm, in particular between 20 and 70 pm, and advantageously have a Span value of between 1 and 3, in particular between 1.5 and 2.

3. Composition according to claim 1 or 2, characterized in that the lipid phase is present at a content of between 0.1% and 5%, in particular between 0.5 and 2%, for example 1%, by volume relative to the total volume of the composition.

4. Composition according to any one of claims 1 to 3, characterized in that the organic particles have an average intensity size less than or equal to 500 nm, in particular between 100 and 300 nm, preferably between 150 and 250 nm, and a polydispersity index typically less than or equal to 0.3, preferably between 0.05 and 0.2, in particular between 0.1 and 0.

15.

5. Composition according to any one of claims 1 to 4, characterized in that the surface agent comprises or consists of chitosan particles.

6. Composition according to claim 5, characterized in that the surface agent comprises or consists of particles of chitosan co-precipitated with an organic acid salt, in particular sodium citrate.

7. Composition according to any one of claims 1 to 6, characterized in that the mass ratio of particulate surfactant / lipid phase L is between 0.1 and 2%.

8. Composition according to any one of claims 1 to 7, characterized in that the aqueous phase Aq is in the form of a gel.

9. Composition according to claim 8, characterized in that the aqueous gel is obtained by gelling an aqueous solution comprising pharmaceutically acceptable viscosifying agents, such as xanthan gum, gellan gum, hyaluronic acid and / or sodium hyaluronate.

10. Composition according to any one of claims 1 to 9, characterized in that said composition comprises at least one active ingredient.

11. Composition according to claim 10, characterized in that the active ingredient is chosen from • prostaglandins, such as latanoprost, travoprost, bimatoprost and tafluprost, possibly in admixture with timolol, levobunolol, dorzolamide and / or brinzolamide, and • steroids, such as hydrocortisone, prednisone, prednisolone, methylprednisolone, betamethasone, dexamethasone, triamcinolone, estradiol, estrone, estriol, progesterone, and mixtures thereof.

12. Ophthalmic composition according to any one of claims 1 to 11 for use as a medicament.

13. Composition for use according to claim 12, wherein the composition is suitable for administration by ocular instillation.

14. Composition for use according to any one of claims 12 and 13, characterized in that: o the composition is useful for the treatment of dry eye, and / or o the liposoluble active ingredient is chosen from prostaglandins, optionally in admixture with timolodol, levobunolol, dorzolamide and / or brinzolamide, and the composition is useful for the treatment or prevention of intraocular hypertension, and / or glaucoma, such as open-angle glaucoma, or o the fat-soluble active ingredient is chosen from steroids, and the composition is useful for preventing, relieving or treating eye inflammations.

15. A process for preparing the composition according to any one of claims 1 to 11, said process comprising the following steps: a) Suspending the particulate surfactant in the aqueous phase Aq, b) Dissolving the liposoluble active ingredient in an oil to form the lipid phase L, c) Addition of the lipid phase L to the suspension obtained in step a), d) Emulsification, so as to obtain an emulsion.

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