IT202000032687A1 - PEDIATRIC OPHTHALMIC COMPOSITION - Google Patents

Description

Description of Patent for Industrial Invention having the title:

?PEDIATRIC OPHTHALMIC COMPOSITION?.

DESCRIPTION

The present invention relates to a pediatric ophthalmic composition for the daily ocular and periocular hygiene of newborns, infants and children. With particular reference to the medical device sector? the use of ophthalmic compositions for the maintenance of ocular hydration and/or to alleviate ailments and discomforts due to allergies, mechanical and environmental stress, dryness, itching and burning of the eye as well as? states of inflammation.

In this regard it is specified that in the context of this discussion with the terms ?ophthalmic compositions? means eye drops, ophthalmic solutions, solutions for ophthalmic use and medical devices such as eye drops.

Anatomically, the eye has an external surface visible to an observer without special instruments and having a substantially spherical shape. The outer surface? posteriorly constituted by the sclera, a thin opaque white tissue, which covers about 95% of the external surface of the eye itself and which is interrupted anteriorly, giving rise to a second spherical portion, the cornea.

The cornea represents the remaining 5% of the external surface of the eye, is renewed every 5-10 days and is ? made up of a single multi-layered biological tissue, transparent to light, devoid of blood vessels and which ? designed to transmit and focus light inside the eye itself and, moreover, to act as a barrier against the entry of bacteria or other pathogens.

The cornea continues in the conjunctiva, a mucous membrane that covers the back face of the two eyelids and which extends from these to the eyeball covering the anterior part around the cornea.

The constituent parts of the external surface of the eye, that is? the cornea and the conjunctiva are previously hydrated by the tear film, a transparent fluid with a mainly lubricating action.

In detail, the tear film plays a fundamental role in maintaining the homeostasis of the external surface of the eye, carrying out various specialized functions aimed at nourishing, lubricating and protecting the external surface itself from bacterial agents.

About this ? good to specify that from an anatomophysiological point of view, to ensure a healthy ocular surface? necessary that the quantity? of the tear fluid is sufficient, the quality? and the composition of the latter respect the physiological parameters and, finally, the blinking mechanism works adequately so? to ensure uniform distribution of the tear fluid itself on the ocular surface.

In fact, tears, in addition to performing an antibacterial function, lubricate the surface of the eye and remove any foreign bodies.

With reference to the pediatric sector, in particular newborns, infants and children, ? the need to formulate new products specifically aimed at the latter in terms of usability? and safety and capable of allowing the removal of secretions produced as a result of inflammatory manifestations (such as blepharitis, conjunctivitis, various types of infections) the consequences of which greatly affect production and quality? of the tear film.

Moreover, ? to be emphasized that the newborn, the infant and the child up to 3 years of age?, are more? adults exposed to bacterial aggression as a cause of their behavior: in fact, they often rub their eyes with their hands after having put them in their mouth and in contact with surfaces of various kinds which very often are not adequately sanitized.

In this scenario, the installation of drops or ointments does not work. easily administered to neonates or infants.

To there? it is added that among the disorders that arise in age? pediatric there ? the failure or insufficient production of tears that pu? cause dry eye syndrome and, if so, is it prolonged over time, it causes damage to both the conjunctiva and the cornea; these tissues being interconnected and externally covering the ocular surface must be considered, and consequently treated, as a ?continuum? and not as independent sections and s? standing.

About this ? good to point out that the? eye ? a sophisticated and complex integrated system exposed daily to insults of various kinds that undermine its stability? and whose correct functionality? it depends on a fragile balance of factors.

The causes of suffering and eye damage are numerous, among the predominant ones are oxidative stress, together with the aforementioned, dry eye syndrome, conjunctivitis, various types of infections and allergic irritations.

In particular, oxidative stress, due to the generation of free radicals, is often linked to photochemical factors or pathological conditions (such as inflammation).

Pi? in detail, the formation of free radicals affects both the ocular surface and the deep tissues; in fact, the light can? reach the macula which represents the portion of the retina where there is the greatest concentration of sensory cells, damaging the pigmented epithelium and the photoreceptors, especially the cones which respond to blue light.

In addition to the aforementioned causes, there are numerous disturbances linked to qualitative and quantitative imbalances of the tear film that afflict a high percentage of the child population.

Furthermore, among the most severe eye allergies there? vernal keratoconjunctivitis, which ? associated with dry eye symptoms in 20% of cases. Ocular inflammation and instability are involved in the pathogenesis of dry eye in atopic and vernal keratoconjunctivitis. of the tear film. Compared to adults, children are more exposed to epithelial damage caused by prolonged inflammation.

In addition, a common cause of ?red eye? ? the activity? sports: swimming in a pool, for example, exposes the eye to contact with water containing chlorine, a highly irritating substance for the conjunctiva. Even excessive exposure to ultraviolet rays from the sun, in the absence of adequate protection, can cause inflammation of the ocular surface. One of the biggest problems common ? linked to exposure in places with a lot of glare (open sea, beach or ski slopes), which can? cause actinic keratoconjunctivitis, an acute inflammation affecting both the conjunctiva and the cornea with symptoms such as watery eyes, pain, eyelid swelling, grittiness in the eyes, blurred or reduced vision.

Among these disturbances is the so-called ?dry eye syndrome? caused by the combination of numerous factors that alter the secretion and/or composition of the tear film or increase its evaporation causing inflammation of the outer surface of the eye.

Pediatric patients suffering from this disorder are often afflicted with burning, itching, difficulty eyelid opening upon awakening, photophobia and visual blurring; is it easy to understand, therefore, how such disturbances can affect the quality? of the patient's life as if left untreated they can cause permanent damage to vision.

Pi? in detail, one of the main causes of the genesis of the condition of dry eye ? associated with the inflammatory state caused by oxygen free radicals.

In fact, recent studies have identified a? High quantity? of these radicals in the tear fluid of pediatric patients affected by dry eye syndrome.

Furthermore, among the many factors that determine the alteration of the secretion and/or composition of the tear film, there are environmental factors such as: air conditioning, heating, excessive exposure to sunlight, wind, salt, smoke and pollution.

To there? it is added that the daily ocular cleansing of the eyelids assumes particular importance in pediatric patients since? the non-removal of eye secretions pu? cause blockage of the tear ducts, causing stagnation of tears in the conjunctival sac and favoring the development of bacterial infections.

In particular, the daily cleaning of the child's and newborn's eyes has an important preventive function against conjunctivitis, by removing the residues of tear secretion.

In fact, in the case of infective conjunctivitis, the aqueous or mucous secretion becomes abundant, especially when the patient awakens.

In this scenario, in parallel with drug therapy, ? Thorough cleaning of the eye and eyelids is essential, removing the secretions several times a day.

Still, the alteration of the tear film can? be caused by particular clinical conditions due to the use of drugs such as, for example, extemporaneous or continuous therapies (in the presence of chronic pathologies), or to allergies which can in turn be the trigger for a severe dry eye condition, such as consequence of the impact of the same on the tear film.

To date, these disturbances are alleviated by the use of traditional ophthalmic compositions generally not aimed at pediatric patients and presenting various drawbacks among which must be counted the fact that they determine a partial and short-term hydration of the external surface of the eye, thus relieving the disturbances described above are only temporary and forcing the user to administer several close doses which in the case of pediatric patients considerably complicate the administration to the latter who become refractory to receiving the therapy.

At the same time, oil-based or gel-based compositions, aimed at increasing the permanence time of the vehicle on the ocular surface, are often the cause of discomfort, blurred vision; to there? it is added that due to their viscosity? result of even more difficult to apply and leading to an interruption of therapy by the patient.

Furthermore, the aforementioned compositions are ineffective in the face of the need to enclose in a single formulation active moisturizing, humectant, antioxidant (and consequently anti-inflammatory) and control of the microbial load, with a protective effect on the ocular surface.

To date, with reference to? Activity? antioxidant, ? It is necessary to respond to these needs by taking food supplements based on vitamins and/or flavonoids which, being absorbed by the gastric and/or intestinal route, are made available to the body systemically, implying the impossibility to mark and maximize the antioxidant effect on the eye.

A further problem? linked to the eye subjected to a bandage that ? more exposed to infectious risk, since gaseous exchanges with the atmosphere are reduced, since the temperature is at the level of the upper ocular surface and, moreover, since the frequency of blinking and tear production is reduced, the microbial load is? also superior.

In addition, the? use more and more? Prevalence of contact lens wear by children and adolescents has increased the incidence of secondary dry eye symptoms. In fact, the use of corneal lenses can increase the symptoms of dry eye.

In pediatrics, dry eye symptoms are associated with infectious and/or inflammatory processes, as occurs in blepharitis on a constitutional basis, where? mainly involved the saprophytic Demodex, with possible bacterial or viral superinfection. Can dry eye be fueled by the concomitant intake of systemic drugs (such as some antihistamines and oral retinoids for the treatment of acne) or by structural alterations affecting the eyelids, caused by chronic inflammation of the eyelid margin. Furthermore, the inflammation/infection of the eyelid margin often also causes the alteration of the composition of the lipid layer, resulting in an altered balance with the other components of the tear film and inducing a more frequent tear film. rapid evaporation of the aqueous component.

A vicious circle is therefore created, fueled by the possible concomitance of alterations of the eyelid conformation, dysfunction of the meibomian glands and styes or chalazi. In these cases, the removal of the eyelid debris and the instillation of the composition object of the present invention make it possible to reduce the associated symptoms, avoid over-infections and facilitate the restoration of the normal functionality of the eyelids. palpebral and glandular. Furthermore, the increasing use of smartphones, PCs and tablets by children and teenagers can be the cause of a dry eye syndrome accompanied by ocular burning, asthenopeic disorders and headache; the symptomatology derives from the very reduced blinking in environments made particularly dry by the presence of heating and air conditioning. About that, ? demonstrated that the blink rate during video game play is one fourth of the rate at rest. A study conducted in 2009 highlighted a statistically significant improvement in acuity? visual interblink after 90 minutes from? Instillation of artificial tears with lubricating and humectant characteristics, demonstrating their effectiveness during activities? visual.

The main task of the present invention ? that of devising a pediatric ophthalmic composition for the daily cleansing of the eyelids and eyes of pediatric patients, specifically newborns, infants and children which is particularly biocompatible and tolerable, and easy to apply.

Another aim of the present invention ? that of devising a pediatric ophthalmic composition able to allow the removal of natural ocular secretions or dirt residues linked to environmental factors or produced following infections or inflammatory manifestations and able to soften the secretions facilitating their detachment and removal through a humectant effect and refreshing.

Another aim of the present invention ? that of devising a pediatric ophthalmic composition which can be used both in normal conditions and in the presence of blepharitis, conjunctivitis and infections of various kinds, alleviating the patient's discomforts.

Another object of the present invention ? that of devising a pediatric ophthalmic composition capable of reducing symptoms such as hyperemia, edema, itching and photophobia, thanks to its decongestant, cleansing and refreshing actions.

Another object of the present invention ? that of devising a pediatric ophthalmic composition which, in the presence of stenosis of the lacrimal ducts, allows the removal of the purulent material produced in the lacrimal sac and which allows to contain the bacterial super-infection due to the stagnation of tears in the conjunctival sac, caused by the obstruction of the nasolacrimal duct.

Another object of the present invention ? that of devising an ophthalmic composition suitable for controlling the microbial load of the eye subjected to bandage in case of amblyopia, and which at the same time has properties? humectants beneficial in case of use of a contact lens. Another object of the present invention ? that of devising a pediatric ophthalmic composition with a washing, refreshing and decongestant action which alleviates the symptoms of allergies and in particular of seasonal allergic conjunctivitis and which allows purulent secretions to be removed and the tear fluid to be diluted, avoiding the stagnation of tears in the conjunctival sac .

A subsequent aim of the present invention ? to devise an ophthalmic composition, presenting activity? lubricant and humectant for cornea and conjunctiva, and having a protective action that allows to increase the stability? of the ocular film for a prolonged period of time, even in the face of unfavorable environmental conditions such as dust, pollen, allergens, mucoid thickeners and salt.

Again, an object of the present invention ? that of devising a pediatric ophthalmic composition suitable for relieving and reducing the symptoms associated with dry eye syndrome and eye fatigue.

Furthermore, another object of the present invention is to devise a pediatric ophthalmic composition with ability? control of the microbial load without being cytotoxic.

Finally, another object of the present invention ? that of devising an ophthalmic composition which allows the aforementioned drawbacks of the prior art to be overcome within the ambit of a simple, rational solution which is easy and effective to use and has a low cost.

Other characteristics and advantages of the present invention will become more evident from the description of a preferred, but not exclusive, embodiment of an ophthalmic composition, particularly antioxidant, anti-allergic and anti-inflammatory.

The above objects are achieved by the present ophthalmic composition having the characteristics of claim 1.

Other characteristics and advantages of the present invention will become more evident from the description of a preferred, but not exclusive, embodiment of a pediatric ophthalmic composition, illustrated by way of example, but not as a limitation, in the attached tables of drawings in which:

figure 1 ? a comparative graph of LDH release expressed in mU/ml. LDH quantified in the negative control (NC) ? was considered as the basal tissue release (55.6 mU/mL);

figure 2 ? a comparative graph of treatment with yellow fluorescent luciferase after 8 hours of treatment. The average value? calculated on a biological triplicate;

figure 3 ? a comparative graph of trans-epithelial electrical resistance (TEER) expressed in ohm*cm<2 > after 8 hours of treatment.

The present invention relates to a pediatric ophthalmic composition.

In detail, with the term ?pediatric? it refers to neonates, infants and children.

It is specified that in the context of this treatment, newborn means the patient from birth up to 28 days of age, infants means the patient from 28 days up to the beginning of weaning and children means patients up to the age of 28 days. onset of puberty.

From an anatomical point of view, it is necessary to underline that the global dimension of the bulb and the axial length undergo a clear change in the first months of life. The axial length of the bulb at birth? between 16.6 and 16.8 mm. The posterior segment of the medulla ? most responsible for these changes, having at birth a size of less than half? compared to the adult. Conversely, the anterior segment, the one directly involved in the dynamics of the medical device, at birth has a size equal to about 75-80% of that of an adult. Therefore, the largest dimensional change ? consequence of the expansion of the scleral surface at the level of the posterior segment of the bulb. Although this development continues until approximately 13 years of age, 50% of the total increase occurs in the first six months of life. In the first 18 months of life, the axial length goes from 16.6 mm to 20.3 mm.

As far as ocular physiology is concerned, at birth the excretory and secretory lacrimal apparatus is functioning in most cases and recent studies show that the lacrimal gland of the newborn is histologically comparable to that of the adult; moreover, 80% of newborns have a normal basal tear secretion within the first two days of life. The reflex tear secretion, on the other hand, normalizes around the second month of life. The analysis of the tear film of the newborn by electrophoresis shows a content of albumin and lysozyme comparable to that of the adult, so as for the other components. The conjunctival sac, sterile at birth, from the fifth day of life shows a bacterial flora comparable to that of the adult. The development of the nasolacrimal duct ? almost complete at birth, even if ? congenital obstruction of the duct is commonly found (in 2-4% of cases) with spontaneous resolution in 75-90% of cases within the first 12 months of life. From 2 to 5 years the axial length of the bulb increases by about 1.1 mm, while between 5 and 13 years there is a slow development with an axial increase of 1.3 mm. As far as the palpebral aperture is concerned, the changes in the vertical dimension of the palpebral fissure are minimal, going from 8.5 mm at one year of life to 9 mm at 10 years of age. The horizontal dimension of the palpebral fissure undergoes rapid growth from the first to the tenth year of life, reaching adult size (18 mm at birth vs 25 mm in the adult). With regard to ocular physiology and ocular adnexa there are no changes with respect to what is described in the previous paragraph for newborns and infants.

The ophthalmic composition includes an aqueous solution, zinc sulfate and D-alpha tocopheryl polyethylene glycol 1000 succinate (Vitamin E TPGS). Preferably, zinc sulphate ? present in a concentration by weight, evaluated with respect to the total weight of the composition, between 0.0001% and 0.01% and the D-alpha tocopheryl polyethylene glycol 1000 succinate ? present in a concentration by weight, evaluated with respect to the total weight of the composition, between 0.001% and 0.1%.

In accordance with a preferred embodiment of the composition, zinc sulfate is present in a concentration by weight, evaluated with respect to the total weight of the composition, between 0.0008% and 0.005% and the D-alpha tocopheryl polyethylene glycol 1000 succinate ? present in a concentration by weight, evaluated with respect to the total weight of the composition, between 0.008% and 0.05%.

Zinc sulphate? an element naturally present in the eye and carries out an activity? Broad spectrum antibacterial and antiseptic.

D-alpha tocopheryl polyethylene glycol 1000 succinate? a very stable form of Vitamin E with good (non-ionic) surfactant action ? adapted to improve the wettability? of the cornea and counteracts tear evaporation. In addition, D-alpha tocopheryl polyethylene glycol 1000 succinate acts as a lubricant, as it? ease? sliding of the eyelid reducing friction and therefore ocular discomfort. Again, D-alpha tocopheryl polyethylene glycol 1000 succinate carries out an antioxidant action as a scavenger of free radicals generated by daily insults to which the eye is exposed. exposed (which cause states of irritation and inflammation). Furthermore, the humectant action of D-alpha tocopheryl polyethylene glycol 1000 succinate protects against dehydration, facilitating healing from infectious states, reducing the inflammatory response caused by hyperosmolarity and tear and lubricating the ocular surface.

Furthermore, the composition includes one or more? distilled water of a plant extract.

Preferably, the aforementioned one or more? Distilled waters include at least one of distilled echinacea water and distilled eyebright water.

Advantageously, the distilled water of echinacea is present in a concentration by weight, evaluated with respect to the total weight of the composition, between 1% and 10%.

Preferably, distilled echinacea water is present in a concentration by weight, evaluated with respect to the total weight of the composition, between 2% and 5%.

Preferably, the distilled water of eyebright ? present in a concentration by weight, evaluated with respect to the total weight of the composition, between 1% and 10%.

Preferably, the distilled water of eyebright ? present in a concentration by weight, evaluated with respect to the total weight of the composition, between 2% and 5%.

The distilled waters used in the formulation are extracted in a steam current and are totally free of preservatives.

Echinacea ? known for its activities antibacterial and antiviral and, applied topically, contributes to the control of ocular bacterial flora thanks to the echinacoside and polyacetylenes.

Euphrasia, on the other hand, is known for its properties astringent and anti-inflammatory attributable to the presence of flavonoids and tannins. Both distilled waters also contribute indirectly, i.e. by increasing the volume and diluting the tear fluid, to reducing the inflammatory state of the ocular surface in case of inflammation of the anterior segment of the eye.

Furthermore, the composition comprises at least one isotonizing agent.

? useful to point out that in the context of this discussion for ?isotonizing agent? means a component suitable for maintaining the osmotic pressure of the present composition at values compatible with the osmotic pressure of the tear film.

In detail, an ophthalmic composition? deemed isotonic when its tonicity? (linked to the permeability? of the cell membrane to solutes) ? equal to that of a 0.9% sodium chloride solution (saline solution) and, therefore, to that of plasma and tear film.

In the present embodiment, the isotonizing agent ? designed to maintain the osmotic pressure substantially equal to 300mOsm/Kg.

Preferably, the isotonizing agent comprises sodium chloride.

The composition comprises at least one carrier agent selected from the list: water for injections and purified water.

Advantageously, the composition comprises at least one buffering agent selected from the list: borate buffer, phosphate buffer, citrate buffer and acetate buffer.

In detail, the aforementioned buffering agent ? citrate buffer.

The citrate buffer? designed to maintain the pH at a value substantially equal to 7.

Advantageously, the composition ? free from preservatives.

Also, the composition? free from the following substances: latex, substances deriving from genetically modified organisms, phthalates, molecules with active? anti-inflammatory and vasoconstrictive which can be identified by the acronym API (active pharmaceutical ingredients), sorbic acid, EDTA and sodium perborate.

? it should be emphasized that the absence of preservatives reduces the risk of adverse effects on the user, such as, for example, eye irritation caused by the use of the preservative itself.

Furthermore, the absence of the preservative allows its use together with other products, eliminating the risk of interaction between the preservative used in the ophthalmic composition and any other drug possibly taken by the patient.

Finally, the? use of substances already? present in the tear fluid prevents any modification of the characteristics of the tear film itself. Finally, the aforementioned composition ? indicated for use in the treatment of pediatric dry eye syndrome, pediatric seasonal allergic conjunctivitis, the symptoms of blepharitis states and for the reduction of ocular inflammatory states of various origins.

SAFETY OF THE FORMULATION FOR PEDIATRIC USE CYTOTOXICITY TESTING?

Literature shows that a concentration of 2.8 mmol/L of Zinc ions ? almost able to halve the population present in a solution of 107 cfu/mL of Staphylococcus aureus microorganism capable of colonizing the eye. This concentration corresponds to 0.08% of zinc sulphate heptahydrate.

However, the aforementioned concentration of zinc sulphate is cytotoxic and would not allow the formulation to obtain a CE marking, especially for use in the pediatric field.

Were therefore carried out various cytotoxicity tests? aimed at determining the quantity? non-cytotoxic concentration of zinc sulphate which can be used in the composition object of the present invention.

The results of these tests are reported below.

TEST 1

Formulation with 0.08% Zinc Sulphate - cytotoxic result according to the UNI EN ISO 10993-5:2009 method.

TEST 2

Formulation containing 0.035% Zinc Sulphate (product ?Zixol?) and subsequent dilutions resulted cytotoxic as such according to the UNI EN ISO 10993-5:2009 method.

TESTS 3

Carried out on the formulation object of the present invention (containing 0.001% Vitamin E TPGS Zinc Sulphate) which resulted non-cytotoxic according to the UNI EN ISO 10993-5:2009 method.

Qualitative evaluation

Degree of cytotoxicity? morphological

Interpretation of results

Acceptability criterion?

Acceptability criterion?

Results

Quantitative evaluation

Density measurements? optics ? OD value

Mean, Standard Deviation and %CV of OD values

% vitality count

Acceptability criterion?

Acceptability criterion?

Acceptability criterion?

Interpretation of results

p

Results

As further confirmation of the safety of the pediatric ophthalmic composition according to the present invention, ? A further study was carried out to ascertain the possible absorption of the product.

CAPACITY ASSESSMENT? OF THE PRODUCT TO MODIFY THE BARRIER FUNCTION OF THE CORNEAL EPITHELIUM

? been studied on an experimental basis the ability? of the composition object of the present invention (COMPOSITION 1 - ACCURA) to modify the barrier function of the corneal epithelium, after repeated applications of the product, using reconstructed human corneal epithelium (HCE = Human Corneal Epithelium). The models of reconstructed human epithelium are very similar in terms of morphology (multi-stratified epithelium), properties and biochemical and physiological properties to living human tissue. The biological relevance and predictivity of these models derive from the presence of a tissue organized in different layers of live cells which allows to evaluate the topical application of products at the real doses and conditions of application. In the study the model of HCE ? treated twice with 30 microL of the solution object of the present invention and with saline solution (negative control) for a total of 8 hours of exposure. Between the two treatments? was an interval of 4 hours defined and before the second exposure, the residual product from the first application? been removed, with the aim of mimicking the physiological clearance of the product from the eye, generally by tearing or blinking.

At the end of the 8-hour exposure, the following endpoints were investigated:

1. (figure 1) Cell membrane damage through the quantification of the release of LDH (Lactate Dehydrogenase) an enzyme which is normally found in the cytosol and which can be measured at the extracellular level only following cellular damage.

2. (figure 2) Integrity? of cellular junctions through the evaluation of the paracellular passage of LY (Lucifer Yellow). In the presence of intact cell junctions, LY has very low permeability, but if the junctions between cells are damaged, LY diffusion increases.

3. (figure 3) Property? of the barrier function through TEER (Trans-Epithelial-Electrical-Resistance). The TEER? the measurement of ion movement across cell junctions regulated by the polarized surface of the cell membrane and the tight cell-to-cell junctions that together impede the movement of solutes and ions across the epithelium. TEER ? then a method of indirect evaluation of the stability? of the cell junctions and consequently a direct measure of the functionality? of the barrier function of the epithelial tissue: TEER reflects the global resistance of the barrier linked to both the structure and the thickness of the epithelium.

COMPOSITION 1:

- Echinacea distilled water 3%

- Distilled water of eyebright 3%

- Zinc sulphate 0.001%

- D-alpha tocopheryl polyethylene glycol 0.01%

- Sodium chloride - at 300 mOsm/kg 0.82%

- Sodium Citrate Trisodium dihydrate 0.35%

- Citric acid monohydrate 0.0032%

- Purified water 94%

The study demonstrated that, after a total of 8 hours of exposure, the formula object of the present invention (COMPOSITION 1 ? ACCURA) did not produce a significant increase in the release of LDH in the extracellular matrix, compared with the negative control; there? does it indicate that the product has not caused damage or induced changes in the integrity? of the cell membrane (Figure 1).

The permeability? of the corneal epithelium, measured by the test with Yellow Lucifer, confirms these results, since? the values of the tissues treated with the formulation object of the present invention do not differ from those treated with the negative control (figure 2).

Furthermore, even the TEER values detected at the end of the exposure do not differ from those relating to the negative control (figure 3).

All of that? confirmation that the product object of the present invention does not induce modifications to the physiology and integrity of the barrier function of the reconstructed human corneal epithelium.

ACTIVITIES CONTAINMENT OF THE MICROBIAL LOAD OF THE PRIVILEGED FORMULATION STUDY 1

? been conducted an experimental study aimed at evaluating the? activity? antimicrobial and/or containment of bacterial proliferation of the preferred formulation object of the present invention against a series of microorganisms commonly present in the ocular microbiome. The ocular surface? continuously exposed to the environment and consequently to different types of microorganisms, starting from birth and throughout life. Many studies have examined the types of microbes that are routinely present on different parts of the ocular surface. Among the types of microorganisms that most? commonly they can be isolated on the conjunctiva, on the eyelids and in tears, staphylococci are found, isolated in all three anatomical compartments, including Staphylococcus aureus. Other species include Streptococcus, Bacillus and Pseudomonas aeruginosa.

A 1988 study describes and compares the typical microorganisms of newborns, born with normal vaginal delivery and cesarean delivery, making a distinction between the characteristic flora of the conjunctival sac on the day of birth and two days after birth: staphylococcus (staphylococcus aureus) are still the most represented microorganisms.

Finally, yes? referring to a publication taken from Current Eye Research which reaffirms that the microorganisms present on the ocular surface come both from the body itself (eyelids, skin, nasopharynx, birth tract) and from the external environment (water, atmosphere, soil ). For the study, the following microorganisms were selected: Pseudomonas aeruginosa and Candida albicans,

In the aforementioned experimental test ? a composition was used comprising (COMPOSITION 1):

- Echinacea distilled water 3%

- Distilled water of eyebright 3%

- Zinc sulphate 0.001%

- D-alpha tocopheryl polyethylene glycol 0.01%

- Sodium chloride at 300 mOsm/kg (0.82%)

- Sodium citrate trisodium dihydrate (0.35%)

- Citric acid monohydrate (0.0032%)

- Purified water 94%

EQUIPMENT

Analytical scale

Vapormatic autoclave

Incubator at 37?C

Incubator at 25?C

Laminar flow hood

Disposable sterile material for microbiology

CULTURE MEDIA AND REAGENTS

Microorganisms:

Pseudomonas aeruginosa ATCC9027, titrated (6.8x10^7/pellet) and dehydrated in pellets, manufacturer Microbiologics;

Candida albicans ATCC 10231, titrated (6.8x10^3/pellet) and dehydrated in pellets, manufacturer Microbiologics;

Triptic Soy Agar Culture Medium: Manufacturer WVR, Product Code 84602.0500 Lot L0289W;

Sabouraud dextrose Agar culture medium: manufacturer WVR, product code 84663.0500 Lot 1443;

Buffer solution pH 7.2 for the preparation of the microbial suspension; Test solution = AccuraBaby solution.

PROCEDURE

Stump preparation

The pellets of the reference strains taken from the refrigerator and acclimatised to room temperature were suitably diluted in a pH 7.2 buffer solution preheated to 37°C. The quantity? initial diluent and the subsequent dilutions used were calculated according to the titre of the various strains, with the aim of reaching a final concentration <100 CFU. Specifically yes? proceeded as follows:

Pseudomonas aeruginosa: the pellet? was transferred to 100 ml of pH 7.2 buffer solution, mixed and transferred to an incubator at 37°C for 1 hour. After incubation? 0.1 ml was taken and transferred to 100 ml of pH 7.2 buffer solution. From this solution, after suitable stirring, ? 1 ml was taken and transferred to 10 ml of pH 7.2 buffer solution. This last solution, with a theoretical microorganism concentration of 68 CFU, is been used for the execution of the test.

Candida albicans: the pellet? was transferred to 10 ml of pH 7.2 buffer solution mixed and transferred to incubator at 37?C for 1 hour. After incubation, 4 ml were taken and transferred to 50 ml of pH 7.2 buffer solution. This last solution, with a theoretical microorganism concentration of 54 CFU, is been used for the execution of the test.

Execution of the test: verification of the reduction and/or containment of the strain title by the sample under test

Strain title:

from the solution of each strain ? 1 mL was transferred in triplicate to 3 Petri dishes.

Test solution (ST):

1 ml of the solution of each strain 1 ml of the sample under examination were transferred in triplicate on three Petri dishes, different for each strain.

Approximately 15 ml of TSA medium prepared in the morning and cooled to 47°C were added to all the plates containing the bacterial strains (Pseudomonas aeruginosa).

Approximately 15 ml of SDA medium prepared in the morning and cooled to 47°C were added to the plates containing the Candida albicans strain.

Simultaneously ? A blank containing only 1 ml of the product to be tested, a process blank containing only the TSA medium and a process blank containing only the SDA medium were performed.

After cooling and solidification, the plates containing the bacterial strains (Pseudomonas aeruginosa) were incubated, upside down, at 37?C -1?C for 24 hours;

the plates containing the Candida albicans strain were incubated, upside down, at 20-25°C for 3 days.

Colony count

At the end of the incubation period (26 hours for the bacterial strains, and 72 hours for the fungal strains), the colonies grown on the single plates were counted.

The following results were obtained:

Number of colonies grown

(CFU/plate)

No colonies grew in the blank plate containing TSA medium only; no colonies grew in the blank plate containing SDA medium only; no colonies have grown in the plate containing the TSA medium test solution; no colonies have grown in the plate containing the SDA medium test solution.

Against Pseudomonas aeruginosa and Candida albicans the ST test solution (COMPOSITION 1) limits the growth of the number of colonies.

STUDY 2

? was conducted a further experimental study aimed at evaluating the activity? antimicrobial of the composition in accordance with the present invention against Staphylococcus Aureus, which ? among the microorganisms commonly found in the ocular microbiome. In particular, the purpose of the study ? been comparing the preferred formulation of the present invention with the performance of two other formulations: a) a variant of the formulation (with only Zinc Sulphate but without Vitamin E TPGS) and b) a product currently on the market (?Zixol?) containing Zinc 0.035% sulfate.

In the aforementioned experimental test ? a composition was used comprising (COMPOSITION 1):

- Echinacea distilled water 3%

- Distilled water of eyebright 3%

- Zinc sulphate 0.001%

- D-alpha tocopheryl polyethylene glycol 0.01%

- Sodium chloride at 300 mOsm/kg 0.82%

- Sodium Citrate Trisodium dihydrate 0.35%

- Citric acid monohydrate 0.0032%

- Purified water 94%

In order to evaluate the? activity? antimicrobial of the composition in accordance with the present invention only the latter? been compared with the following compositions.

EVALUATION OF THE ACTIVITY? ANTIMICROBIAL OF THE COMPOSITION IN ACCORDANCE WITH THE PRESENT INVENTION TOWARDS STAPHYLOCOCCUS AUREUS SAMPLES USED

EQUIPMENT:

- Analytical scale

- Vapormatic autoclave

- Incubator at 37?C

- Laminar flow hood

- Disposable sterile material for microbiology

CULTURE MEDIA AND REAGENTS

- Microorganism Staphylococcus aureus ATCC 6538 titrated (5.3x10^3 per pellet) and dehydrated in pellets, manufacturer Microbiologics;

- Petri dishes of 90 mm in diameter;

- Triptic Soy Agar culture medium: manufacturer VWR, product code 84602.0500 Lot M0079W, expiry 03/30/2024;

- pH 7.2 buffer solution for the preparation of the microbial suspension;

PROCEDURE

Stump preparation

The pellet of the reference strain acclimatised to room temperature? was transferred into a sterile container containing 50 ml of pH 7.2 buffer solution preheated to 37?C.

After thorough mixing, the tube with the pellet ? was transferred to an incubator at 37°C for 1 hour (STRAIN solution with a theoretical concentration of about 106 CFU).

Test execution

Strain title:

- 1 ml of the STRAIN solution ? was transferred in triplicate to 3 Petri dishes. Test solution 1:

- 1 ml of STRAIN solution 1 ml of ST1 solution were transferred in triplicate on three Petri dishes.

Test Solution 2:

- 1 ml of STRAIN solution 1 ml of ST2 solution were transferred in triplicate on three Petri dishes.

Test Solution 3:

- 1 ml of STRAIN solution 1 ml of ST3 solution were transferred in triplicate on three Petri dishes.

Approximately 15 ml of TSA medium cooled to 47°C was added to all plates.

Simultaneously ? A blank was performed containing only 1 ml of the product to be tested and a process blank containing only the TSA medium. After cooling and solidification all the plates were incubated, upside down, at 37°C ?1°C for 24 hours.

Colony count

At the end of the incubation period (26 hours) the colonies grown on the single plates were counted.

The following results were obtained:

No colonies grew on the plates containing the blanks of the three solutions and the process blank.

In the plates containing the ST1, ST2, ST3 STRAIN solution, it is noted that in the ST1 STRAIN solution (COMPOSITION 1) the growth of the number of colonies ? lower than the other two solutions.

In a second aspect, the present invention relates to a pediatric ophthalmic formulation, comprising the previously described composition and a container containing the latter and provided with a dispensing mouth suitable for dispensing the composition in the form of a spray. Preferably, the container? of the type of a polydose.

In addition, the container ? free from preservatives.

The spout? configured to allow atomization of the composition in accordance with the present invention.

There? it allows the composition according to the present invention to be dispensed effectively and accurately, preventing the patient from being deprived of administering the latter when moving.

Below ? reported a study of stability? of the formulation in accordance with the present invention.

STABILITY? OF THE OBJECT FORMULATION OF THE PRESENT INVENTION

? Has a stability study been conducted? in ICH storage conditions according to the ICH Q1A guideline of February 2003 Stability Testing Guidelines: stability testing of new drug substances and products with the aim of obtaining experimental data to support the period of validity? of the proposed product and of the relative modalities? of conservation. Specifically, the design of the stability study? ? was the following: a) study of the stability? LONG TERM, at 25?C 2?C/60%UR 5% (long term storage condition climatic zones I and II) for 36 months; and b) study of stability? ACCELERATED (accelerated conditions) at 40?C 2?C /75%UR 5% for 12 months (worst case).

The solution object of the present invention ? was subjected to sterilization by filtration and then divided into single-dose strips of PELD (low density polyethylene: PE/LD Purell PE 1840 H), of 5 vials of 0.5 mL each, sterile by irradiation with 25 kGy gamma rays.

The single-dose container? was chosen as the case pi? challenger because the thickness of the walls and the density? of the material are less than the polydose container.

Here are the reference specifications:

Appearance control ? been carried out for visual observation, the controls of pH and osmolality? were carried out according to Ph.Eur c.e. respectively using pH meter and osmometer. The sterility? ? tested according to Ph.Eur. there is.

Below are the results of the study that confirm the stability of the formulation object of the present invention.

Table 1 ? Accelerated conditions

Table 2 ? Long term conditions

Yup ? in practice it has been observed how the described invention achieves the proposed aim and objects.

It should be emphasized that the particular precaution of providing for the synergistic combination of zinc sulphate and D-alpha tocopheryl polyethylene glycol 1000 succinate (Vitamin E TPGS) allows to carry out activities antimicrobial against Staphylococcus aureus.

Specifically, the synergistic combination of 0.001% Zinc Sulphate and D-alpha tocopheryl polyethylene glycol 1000 succinate performs the same effectiveness as a solution where the Zinc Sulphate ? present at 0.035% by carrying out, however, activities? cytotoxic.

Again, the formulation that includes the synergistic combination of zinc sulfate and D-alpha tocopheryl polyethylene glycol 1000 succinate, equal? of percentage concentration, shows improved results compared to formulations containing only zinc sulphate in relation to the activity? antimicrobial against Staphylococcus aureus.

To there? it is added that the ophthalmic composition in accordance with the present invention performs diluting, refreshing and decongestant, allowing to increase the volume of tears.

Furthermore, the synergistic combination of zinc sulphate and D-alpha tocopheryl polyethylene glycol 1000 succinate (Vitamin E TPGS) allows effective cleansing of the eyelids and eyes of newborns, infants and children together with wetting of the ocular surface as an alternative to conventional saline which perform a mere washing action, facilitating the removal of natural eye secretions or dirt residues due to environmental factors such as dust, pollen, allergens, mucoid thickeners, saltiness or produced following infections or inflammatory manifestations, including conjunctivitis and blepharitis of bacterial etiology.

Again, D-alpha tocopheryl polyethylene glycol 1000 succinate carries out activity antioxidant following the hydrolysis of the ester bond of the d-alpha tocopherol segment with PEG 1000. In vivo the predominant reaction is? the enzymatic hydrolysis that takes place inside the cells by cytoplasmic esterases. These release D-alpha tocopheryl polyethylene glycol 1000 succinate which, localizing at the level of the cell membrane, performs an antioxidant function, protecting the membrane from lipid peroxidation and counteracting its damage.

In addition, in the event of stenosis of the tear ducts, the composition according to the present invention allows both to remove purulent secretions and to dilute the tear fluid itself, also decreasing greatly increased risk of bacterial infections. Indeed, the obstruction of the nasolacrimal duct causes the stagnation of tears in the conjunctival sac favoring the development of bacterial infections.

Even in the case of amblyopia, the composition in accordance with the present invention allows to control the microbial load of the eye subjected to bandaging thanks to the dilution of the microorganisms due to the increase in tear volume and to a rapid washing and decongestant action.

Furthermore, the activity decongestant and refreshing of the present composition counteracts the symptoms caused by nocturnal ocular edema. In addition, the composition object of the present invention ? indicated in case of immune response with pro-inflammatory pictures and hypersensitivity reaction, which are often found in allergic children and who use corneal lenses.

To there? it is added that the ophthalmic composition object of the present invention ? It has been specifically formulated with highly biocompatible ingredients to ensure tolerance, allowing to obtain a sterile, isotonic, physiological pH, non-viscous and preservative-free saline composition.

Furthermore, the aforementioned synergistic combination of components allows to alleviate the discomfort caused by a seasonal allergic rhinoconjunctivitis, decreasing the sensation of occasional dry eye or deriving from a blepharitis in progress or following pharmacological therapy.

The lubricating, washing and humectant action of the present composition allows to remove debris from the lens, remove corneal catabolites, rehydrate the lens and reduce the symptoms associated with the use of contact lenses. In the event that a contact lens is worn continuously, the use of this composition upon waking hydrates the lens and avoids dangerous phenomena of epithelial adhesion, especially in the presence of lenses for aphakia. Furthermore, in the case of inflammatory states or mild infections for which pharmacological treatment is not foreseen, the present composition reduces the associated symptoms, the inflammatory state, cleanses and removes secretions and ocular and eyelid debris and avoids over-infections; in fact, by diluting the pro-inflammatory substances and reducing eyelid friction and the bacterial load, it facilitates the action of the local immune defenses and spontaneous healing.

Finally, the composition according to the present invention performs an action of control and containment of the microbial load present in the ocular microbiome.

Claims (11)

1) Pediatric ophthalmic composition, comprising an aqueous solution, zinc sulfate and D-alpha tocopheryl polyethylene glycol 1000 succinate (Vitamin E TPGS). 2) Composition according to claim 1, characterized in that it comprises one or more? distilled waters of a plant extract. 3) Composition according to one or more? of the preceding claims, characterized by the fact that said one or more? Distilled waters include at least one of distilled echinacea water and distilled eyebright water. 4) Composition according to one or more? of the preceding claims, characterized in that it comprises at least one osmotic agent capable of maintaining the osmotic pressure substantially equal to 300mOsm/Kg. 5) Composition according to one or more? of the preceding claims, characterized in that said osmotic agent comprises sodium chloride. 6) Composition according to one or more? of the preceding claims, characterized in that it comprises at least one buffering agent selected from the list: borate buffer, phosphate buffer, citrate buffer and acetate buffer. 7) Composition according to one or more? of the preceding claims, characterized in that it comprises purified water. 8) Composition according to one or more? of the preceding claims, characterized in that said zinc sulphate is present in a concentration by weight, evaluated with respect to the total weight of the composition, comprised between 0.0001% and 0.01%, and said D-alpha tocopheryl polyethylene glycol 1000 succinate ? present in a concentration by weight, evaluated with respect to the total weight of the composition, between 0.001% and 0.1%. 9) Composition according to one or more? of the preceding claims, characterized by the fact that ? free of: genetically modified organisms, phthalates, substances with active? anti-inflammatory, substances with activity? vasoconstrictive, pharmaceutical active ingredients, sorbic acid, EDTA, sodium perborate. 10) Composition according to one or more? of the preceding claims, for the treatment of pediatric dry eye syndrome, for the alleviation of irritative states due to pediatric seasonal allergic conjunctivitis and the symptoms of blepharitis states and for the reduction of ocular inflammatory states. 11) Pediatric ophthalmic formulation, comprising the composition according to one or more of the preceding claims and a container containing said composition and provided with a dispensing mouth suitable for dispensing said composition in the form of a spray.