A topical composition designed to maintain and / or restore the integrity of mucous membranes and damaged epidermis

Abstract

Disclosed is a topical mucoadhesive composition comprising hyaluronic acid or a salt thereof, choline alfosserate, an ascorbic acid ester at a concentration ranging from 0.050% w / w to 0.0004% w / w, and at least one pharmaceutically acceptable excipient or carrier.

Description

Technical Field

[0001] The present invention relates to a topical composition for maintaining and / or restoring the integrity of damaged mucous membranes and epidermis.

Background Art

[0002] The integrity of skin tissue and mucous membranes can be affected by various exogenous and endogenous causes such as vitamin deficiencies, incorrect diet, poor hygiene, bacterial, viral or fungal infections, intestinal dysbiosis, alterations of the mucosal microbial flora, endocrine imbalances, debilitating diseases, genetic factors, mechanical, physical, chemical and traumatic factors, radiation, etc.

[0003] Patent Document 1 (EP2646036) Collinear Phosphoceramide discloses a composition useful for maintaining and restoring the integrity of mucous membranes and epidermis containing and hyaluronic acid. Collinear Phosphoceramide is known as a nootropic substance that can improve the nutrition of brain cells (activate blood supply and cell metabolism), and as a result, improve intellectual function.

[0004] Patent Document 2 (WO93 / 19730) Collinear Phosphoceramide discloses its use for skin and hair, which has moisturizing, emollient, elasticising, repairing, and volume-enhancing actions.

[0005] As disclosed in Patent Document 2 (WO93 / 19730), Collinear Phosphoceramide is substantially non-toxic systemically and has significant topical tolerability. In the pharmaceutical field, it is known for use in injectable and oral compositions for treating alterations of the cognitive functions, and as a growth hormone secretagogue.

[0006] Hyaluronic acid is widely present in various body tissues of humans and animals, and has high viscoelasticity and the ability to retain water up to 1000 times the weight of hyaluronic acid. Mainly in the form of sodium hyaluronate, it is widely used in the formulations of cosmetics and pharmaceuticals for mucoadhesive, anti-inflammatory, and reparative purposes.

[0007] Ascorbyl palmitate (AP), a liposoluble form of vitamin C, is effective in protecting lipids from peroxidation and is mainly used in cosmetics at a concentration in the range of 1.0 - 0.1% w / w for antioxidant and anti-radical purposes in anti-aging treatments. Ascorbyl palmitate, like lipids, easily penetrates the skin and mucous membranes, exhibits antioxidant effects, and prevents cell aging and collagen denaturation. In addition, ascorbyl palmitate is sold in the form of dietary supplements for the purpose of preventing the onset of joint pain and rheumatism by promoting iron absorption, assisting blood circulation, and promoting collagen production. Also, ascorbyl palmitate is used as a food additive (E304) for antioxidant purposes. Furthermore, ascorbic acid and its esters, especially L-ascorbyl palmitate, are known to have significant inhibitory activity against hyaluronidases, enzymes responsible for the degradation of hyaluronic acid.

[0008] There is a particular need for a composition that is useful for maintaining and restoring the integrity of the mucous membrane, adaptable to various types of formulations for different application sites, and having a synergistic effect between components and an improved stability against hyaluronidase.

Prior Art Documents

Patent Document

[0009]

Patent Document 1

Patent Document 2

Summary of the Invention

[0010] Currently, the topical use of a combination of high molecular weight hyaluronic acid, ascorbic acid, preferably at a concentration of 0.05% w / w or less of ascorbyl palmitate, and a bio / mucoadhesive matrix on damaged mucous membranes and epidermis has been found to have superior efficacy and flexibility of application and to maintain and restore the integrity of mucous membrane and epidermal tissues compared to known formulations. Collinear Phosphoceramide The combination of the present invention is particularly useful for the treatment of irritative / inflammatory conditions (such as burns, ulcers, heat rashes, dermatitis, etc.) of damaged mucous membranes and epidermis.

[0011] The combination of the present invention is particularly useful for the treatment of irritative / inflammatory conditions (such as burns, ulcers, heat rashes, dermatitis, etc.) of damaged mucous membranes and epidermis. Therefore, the present invention provides a topical mucoadhesive composition comprising hyaluronic acid or a salt thereof, Collinear Phosphoceramide an ascorbic acid ester at a concentration in the range of 0.050% w / w to 0.0004% w / w, and at least one pharmaceutically acceptable excipient or carrier.

[0012] Ascorbic acid esters, preferably ascorbyl palmitate, are present in the formulations of the present invention at a lower concentration than those conventionally used in cosmetics and skin preparations, usually as antioxidants and radical scavengers (0.500 - 0.100% w / w).

[0013] The best balance between the anti - hyaluronidase effect and the mucoadhesiveness of the formulation is considered to be achieved at the indicated concentration of the ascorbic acid ester.

[0014] The average molecular weight of hyaluronic acid is preferably from 800,000 to 4,000,000 Da.

[0015] Hyaluronic acid can be present in the formulation in the form of a sodium salt at a concentration in the range of 20.000% w / w to 0.0005% w / w.

[0016] Collinear Phosphoceramide is present at a concentration in the range of 0.0005 to 10.000% w / w, preferably 0.001% to 1.000% w / w.

[0017] The composition of the present invention comprises one or more mucoadhesive agents selected from β-glucan, sodium carboxymethyl β-glucan, chitosan, carboxymethyl chitosan, carboxymethyl cellulose, hydroxyethyl cellulose, carbomer, PVA (polyvinyl alcohol), PVP (polyvinylpyrrolidone), polycarbophil (polyacrylic acid crosslinked with divinyl glycol), PVM / MA copolymer (copolymer of methyl vinyl ether and maleic anhydride), and VP / eicosene copolymer (1-vinyl-2-pyrrolidinone-1-eicosene (1:1)).

[0018] According to a further aspect, the composition of the present invention can also contain further active ingredients useful for the topical treatment of the mucosa, such as those described in Martindale, The Complete Drug Reference, 34th Edition.

[0019] The composition of the present invention can be formulated by a method suitable for topical administration according to well-known conventional methods as described in Remington, The Science and Practice of Pharmacy, 20th Edition.

[0020] Known excipients or carriers, such as film formers, those described in the Handbook of Pharmaceutical Excipients, 6th Edition, Pharmaceutical Press, can also be added to optimize the specific use of the composition.

[0021] Examples of preferred formulations are gels, oil-in-water (o / w) and water-in-oil (w / o) emulsions, creams, ointments, sprays, powders, lotions and foams.

[0022] Particularly preferred compositions are in the form of aqueous gels (hydrogels), which can be obtained by using pharmaceutically acceptable polymers that can absorb a significant amount of water and thus adhere to the mucosa (mucoadhesion).

[0023] The mucoadhesion of the composition of the present invention ensures a sufficient residence time on the mucosa, which is subject to the leaching action of physical and mechanical factors that can reduce the residence time of the active ingredient, for example in the case of the oral mucosa.

[0024] The composition according to the present invention is suitable for human or veterinary use, and combines the anti-inflammatory and tissue repair effects of hyaluronic acid, Collinear Phosphoceramide the film-forming and protective effects of the bio / mucoadhesive formulation matrix containing, and the anti-hyaluronidase effect of low-concentration ascorbyl palmitate.

[0025] In a preferred embodiment, hyaluronic acid and an ascorbic acid ester, preferably ascorbyl palmitate, are in a molar ratio in the range of 60:1 to 1:1 (relative to the disaccharide units of hyaluronic acid). More preferably, hyaluronic acid and the ascorbic acid ester are in a molar ratio in the range of 20:1 to 1:1, and the best anti-hyaluronidase activity is obtained at said ratio.

[0026] In fact, in the presence of hyaluronidase, hyaluronic acid rapidly degrades to 95 - 97% after only 5 hours.

[0027] As seen in the following examples, ascorbyl palmitate has been proven to be remarkably effective in slowing down the enzymatic reaction rate and consequently reducing the undesirable degradation of hyaluronic acid.

[0028] In fact, after 5 hours, the composition containing ascorbyl palmitate showed up to 50% less HA degradation than that observed in samples used alone or in combination with Collinear Phosphoceramide hyaluronic acid.

[0029] After 24 hours, the composition containing ascorbyl palmitate showed up to 27% less HA degradation than that observed in samples used alone or in combination with Collinear Phosphoceramide hyaluronic acid.

[0030] The anti - hyaluronidase activity of ascorbyl palmitate acts synergistically with the action of hyaluronic acid, whether endogenous or exogenous to the tissue.

[0031] The anti - hyaluronidase action increases the anti - inflammatory and tissue repair effects of hyaluronic acid, protects hyaluronic acid from the enzymatic action of endogenous and bacterial hyaluronidases, ensures a lengthy functional residence in situ, and is thus of utmost importance, along with greater bioavailability and structural integrity.

[0032] Furthermore, the anti - hyaluronidase action also has an anti - bacterial / anti - inflammatory effect. In fact, many bacteria exert an inflammatory effect by producing specific hyaluronidases that promote tissue colonization, thereby facilitating the onset and spread of bacterial infection / contamination at the local level.

[0033] Based on the above considerations, the anti-inflammatory / tissue repair effect of HA is protected and strongly enhanced by its anti-hyaluronidase effect and the presence of a film-forming and bio / mucoadhesive matrix that can form an environment suitable for tissue protection and repair (wound healing).

[0034] The composition of the present invention can be applied to external mucosae such as the mouth and the entire oral mucosa, nasal mucosa, ocular mucosa, auricular mucosa, genital mucosa, anal and rectal mucosa.

[0035] The composition of the present invention is useful for the prevention and treatment of inflammatory diseases and / or those accompanied by damage to the oral mucosa, the prevention and / or treatment of damaged and / or inflamed gums, and the treatment of radiation-induced mucositis.

[0036] Inflammation and lesions of the oral mucosa mean, for example, gingivitis, mucositis (including oral ulcers, recurrent oral ulcers), stomatitis, glossitis, etc.

[0037] These diseases may have different etiologies, for example, they may have mechanical, chemical or pathological causes (infections, oral hypoplasia or intestinal hypoplasia).

Examples

[0038] The following examples further illustrate the present invention. Percentages are expressed as parts by weight (w / w) relative to the total amount of the composition.

[0039] Example 1 - Liquid Composition for Oral Mucosa

Table A

[0040] Example 2 - Gel Composition for Treating Oral Ulcers

Table B

[0041] Example 3 - Gel Composition for Treating Oral Mucositis

Table C

[0042] Example 4 - Liquid Composition for Treating Oral Mucositis

Table D

[0043] Example 5 - Gel Composition for Use in Nasal Mucosa

Table E

[0044] Example 6 - Liquid Composition for Use in Ocular Mucosa

Table F

[0045] Example 7 - Liquid Composition for Use in Aural Mucosa

Table G

[0046] Example 8 - Liquid Composition for Use in Vaginal and Vulvar Mucosa

Table H

[0047] Example 9 - Gel Composition for Use in Vaginal and Vulvar Mucosa

Table I

[0048] Example 10 - Gel Composition for Use in Male Genital Mucosa

Table J

[0049] Example 11 - Gel Composition for Use in Anal and Rectal Mucosa

Table K

[0050] Example 12 - Gel Composition for Treating Inflammatory Gingiva

Table L

[0051] Example 13 - Gel Composition for Treating Damaged Gingiva

Table M

[0052] Example 14 - Vaginal Pessary

Table N

[0053] Example 15 - Enema Composition for Anus

Table O

[0054] Example 16 - Study on Hyaluronidase Degradation in Formulations Containing Glycerophosphocholine and Ascorbyl Palmitate The purpose of this study was to evaluate the enzymatic degradation of two different batches of hyaluronic acid (HA) containing different materials in formulations by hyaluronidase, an enzyme family that catalyzes the degradation of this polysaccharide. The molecular weight distribution of the degraded acid is analyzed using high performance size exclusion chromatography coupled with a Triple Detector Array (HP-SEC-TDA).

[0055] The reagents and materials used were as follows. Hyaluronic acid (Mw 1348000 Da), Contipro a.s. (Czech Republic) Hyaluronic acid (Mw 1450000 Da), Lehvoss Italia S.r.l. (Italy) Glycerophosphocholine Ascorbyl palmitate Citric acid monohydrate, Sigma Aldrich Deionized H2O obtained with a Culligan apparatus Hyaluronidase from bovine testis type I-S, 400 - 1000 units / mg, Sigma Aldrich NaNO3, Sigma Aldrich NaN3, Sigma Aldrich Snap-it vial 0.3ml, 11.6x32mm, Thermo Scientific Snap-it cap 11mm, Thermo Scientific Sodium dihydrogen citrate, Sigma Aldrich

[0056] Prepared and Tested Samples HA Contipro (Solution A): Approximately 50 mg of HA sample was dissolved in 50 ml of citrate buffer (pH = 6.06) and stirred at 38 °C for 16 hours. The concentration of the hyaluronic acid solution is approximately 1 mg / ml. 10 ml of the solution was hydrolyzed. HA Levhoss (Solution B): Approximately 50 mg of HA sample was dissolved in 50 ml of citrate buffer (pH = 6.06) and stirred at 38 °C for 16 hours. The concentration of the hyaluronic acid solution is approximately 1 mg / ml. 10 ml of the solution was hydrolyzed. HA Contipro + GPC (Solution C): 7.5 mg of glycerophosphocholine was added to 10 ml of Solution A. HA Levhoss + GPC (Solution D): 7.5 mg of glycerophosphocholine was added to 10 ml of Solution B. HA Contipro + GPC + AP (Solution E): 5 mg of ascorbyl palmitate and 7.5 mg of glycerophosphocholine were added to 10 ml of Solution A. HA Levhoss + GPC + AP (Solution F): 5 mg of ascorbyl palmitate and 7.5 mg of glycerophosphocholine were added to 10 ml of Solution B.

[0057] Preparation of Buffer Solution Solution 1: 420 mg of citric acid monohydrate was dissolved in 100 ml of deionized water at room temperature for 30 minutes (for 30'). Solution 2: 9.212 g of sodium citrate dihydrate was dissolved in 100 ml of deionized water at room temperature for 30 minutes (for 30'). Buffer solution: Solution 1 was added to Solution 2 and stirred at room temperature for 30 minutes (for 30'). As a result, pH = 6.06.

[0058] Preparation of enzyme solution Hyaluronidase: Approximately 10 mg of hyaluronidase was dissolved in approximately 1 ml of H2O and stirred at room temperature for 3 hours (10 mg / ml).

[0059] Enzymatic degradation procedure: Before adding 67 μl of the enzyme solution (HA / hyaluronidase 15 / 1 w / w), the solution was stirred in an oil bath at 38 °C for approximately 1 hour. Aliquots of the solution were taken at different times (1 hour; 2 hours; 3 hours; 4 hours; 5 hours; 6 hours, 24 hours) and heated at 100 °C for 5 minutes (for 5') using a thermal shaker to denature the enzyme. The solution was filtered to remove the precipitated enzyme (LLG-Syringe filter CA pore size 0.20 μm. Φ 13 mm). Before HP-SEC-TDA analysis, the solution was diluted to 0.5 mg / ml of HA with the mobile phase.

[0060] HP-SEC-TDA analysis Chromatographic Conditions Two equivalent HP-SEC-TDA apparatuses were used. 1) The Viscotek system model TDA305 equipped with a refractive index, right and low angle light scattering, and viscometer detectors was used under the following conditions. Column: Two columns of TSKGMPWXL 13 μm. 7 mm ID x 30 cm L. Tosoh Bioscience; Mobile phase: 0.1 M NaNO3 + 0.05% NaN3; Injection volume: 100 μl; Temperature: 40 °C; Flow rate: 0.6 ml / min. 2) The OmniSEC Multi-detector System (Malvern Instruments Ltd. UK) equipped with a refractive index, right and low-angle light scattering, and viscometer detector was used under the following conditions. Column: Two columns of TSKGMPWXL 13 μm. 7 mm ID x 30 cm L Tosoh Bioscience; Mobile phase: 0.1 M NaNO3 + 0.05% NaN3; Injection volume: 100 μl; Temperature: 40 °C; Flow rate: 0.6 ml / min. The system was calibrated with pullulan standards for molecular weight, polydispersity index, and intrinsic viscosity certification (PolyCAL-PullulanSTD-Malvern Panalytical).

[0061] Chromatogram refinement Chromatograms from TDA305 were acquired and refined using OmniSEC software version 4.6.2, while chromatograms from the OmniSEC multi-detector system were acquired and refined using OmniSEC software version 10.10.

[0062] Results Molecular Weight Distribution Evaluation Initial sample preparation Approximately 50 mg of hyaluronic acid was dissolved in 50 ml of citrate buffer, stirred for 16 hours, diluted to 0.5 mg / ml with the mobile phase, and injected once. To evaluate the molecular weight distribution of hyaluronic acid, the 0.155 dn / dc value known in the literature (see Theisen A. Johann C. Deacon MP. Harding SE. Refractive Increment Data-Book. Nottingham University Press. 2000) was used. The HA sample has an elution time of 10 - 15 ml, a large bell-shaped chromatographic peak, and a very high polydispersion index value. After the HA peak, the mobile phase profile elutes. Table 1 reports the analysis results regarding the weight-average molecular weight (Mw Da), number-average molecular weight (Mn Da), peak molecular weight (Mp Da), and polydispersity index (Mw / Mn), and further reports the intrinsic viscosity value ([η] dl / g), hydrodynamic radius (Rh nm), and recovery rate % by the TDA technique. The values of a and logk representing the slope and intercept constants of the Mark-Houwink curve were also reported. The results are the average of two different analyses, and the values of Mn and Mw are rounded to thousands of Da.

[0063]

Table 1

[0064] Enzymatic Hydrolysis Sample The hyaluronidase action on hyaluronic acid has been widely studied and reported. To study the enzyme activity of hyaluronidase on the prepared sample, the kinetics of hydrolysis was monitored by the HP-SEC-TDA method. The percentage decrease in the molecular weight (in terms of Mw) of the polysaccharide component was determined using the following formula.

[0065]

Number

[0066] HP-SEC-TDA HA Degradation by Hyaluronidase The sample was hydrolyzed twice to confirm the reproducibility. In the following table, the average value of the molecular weight (rounded to the nearest percentile) of both experiments and the corresponding reduction rate are reported.

[0067] [Table 2]

[0068] [Table 3]

[0069] [Table 4]

[0070] [Table 5]

[0071] [Table 6]

[0072] [Table 7]

[0073] Conclusion Such hyaluronic acid samples are all rapidly degraded to 95 - 97% in 5 hours in the presence of hyaluronidase. The hyaluronic acid manufactured by Lehvoss is degraded slightly faster than that manufactured by Contipro, even if the decrease in Mw is exactly the same after 24 hours. Glycerophosphocholine does not contribute to the enzyme reaction rate, as can be seen from the results of Solution C and Solution D. Conversely, the addition of ascorbyl palmitate has been shown to significantly reduce the enzyme reaction rate and undesired HA degradation, which is effectively desirable. In fact, after 5 hours, the HA degradation rate of each of Solutions A to D was up to 97%, while Solutions E and F, which feature ascorbyl palmitate, had HA degradation rates of 49% and 62% respectively (consistent with the differences in degradation rates observed in HA formulations such as Solutions A and B). This represents an improvement of up to 50% over solutions without ascorbyl palmitate. After 24 hours, Solutions E and F, which feature ascorbyl palmitate, showed HA degradation of 72% and 77% respectively. This represents an improvement of up to 27% over solutions without ascorbyl palmitate.

Claims

1. A topical mucoadhesive composition comprising hyaluronic acid or a salt thereof, choline alfoscerate, ascorbyl palmitate at a concentration in the range of 0.050% w / w to 0.0004% w / w, and at least one pharmaceutically acceptable excipient or carrier

2. The composition according to claim 1, wherein the hyaluronic acid or a salt thereof has a molecular weight in the range of 800,000 to 4,000,000 Da.

3. The composition according to claim 2, wherein the hyaluronic acid is in the form of a sodium salt and its concentration is in the range of 20.000% w / w to 0.0005% w / w.

4. The composition according to any one of claims 1 to 3, wherein the concentration of the choline alfoscerate is 0.0005% w / w to 10.000% w / w.

5. The pharmaceutically acceptable excipient or carrier is selected from β-glucan, sodium carboxymethyl β-glucan, chitosan, carboxymethyl chitosan, carboxymethyl cellulose, hydroxyethyl cellulose, carbomer, polyvinyl alcohol, polyvinylpyrrolidone, polycarbophil, PVM / MA copolymer (copolymer of methyl vinyl ether and maleic anhydride), and VP / eicosene copolymer (1-vinyl-2-pyrrolidinone-1-eicosene (1:1)), the composition according to any one of claims 1 to 4.

6. The composition according to any one of claims 1 to 5, in the form of a hydrogel, lipogel, anhydrous gel, O / W or W / O emulsion, solution or suspension.

7. For topical use for maintaining and restoring the integrity of mucous membranes and skin, and for preventing and treating inflammatory diseases affecting them, the composition according to any one of claims 1 to 5.

8. The composition for use according to claim 7, wherein the mucous membranes include the mucous membranes of the oral cavity, gums, nose, eyes, ears, vagina, vulva, anorectal and genital areas.

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