Composition for cosmetic treatment of the dermis

Abstract

A composition for cosmetic treatment of the dermis comprises: a) polycaprolactone, b) b) a recombinant protein of type I collagen and c) an admixture of amino acids formed by glycine, valine, proline and alanine, wherein, with respect to the total weight of said admixture of amino acids, the fraction by weight of glycine is between 25% and 40%, the fraction by weight of valine is between 15% and 25%, the fraction by weight of proline is between 10% and 20% and the fraction by weight of alanine is between 20% and 35%.

Description

[0001] COMPOSITION FOR. COSMETIC TREATMENT OF THE DERMIS

[0002] Technical field

[0003] The present invention relates to a composition for the cosmetic treatment of the dermis, particularly in the treatment of skin blemishes as well as its use for skin improvement.

[0004] Technological background

[0005] Skin ageing is a common dermatological problem resulting both from intrinsic processes, which arise from physiological changes influenced by genetic and hormonal factors, and from extrinsic processes, linked to environmental factors that lead to histological changes, due for instance to sun exposure, cold or chemical agents of various kinds. Skin ageing is mainly manifested by a loss of elasticity and the appearance of wrinkles, folds and / or skin blemishes, which are generally counteracted by applying creams, serums, treatments or by subcutaneous injection of fillers that can relax the skin and make it more elastic and firm.

[0006] Some of the most popular technigues involve introducing a dermal filler, often based on hyaluronic acid, by means of microinjection at the depressions and / or sagging skin to be treated : in this way, the skin is plumped up and the skin tissue is more relaxed and toned. Among the fillers available are so-called "permanent fillers", i.e. substances that, after being injected into the dermis, are not reabsorbed and remain in the body; however, this causes a potential risk of allergic and / or inflammatory reactions, due to the poor bio-compatibility of the injected substances. To date, in fact, their use has been largely superseded by reabsorbable fillers, which are, in fact, absorbed by the patient's body, by which they are better tolerated, guaranteeing greater safety.

[0007] Although technigues involving the use of dermal fillers are widespread and popular, reabsorbable fillers have a temporary effect and often hydrolyse rather guickly, thus reguiring freguent reapplication to maintain their effect on the skin.

[0008] Alternative technigues to the use of fillers are bio-revitalising substances, which aim to stimulate the endogenous production of structural protein elements intended to form a solid and stable network capable of supporting the epidermis, ensuring tone and firmness.

[0009] Although appreciated, known bio-revitalising substances have some aspects for improvement. Indeed, there is a need to design and produce injectable compositions that are increasingly effective in stimulating the natural production of the skin's structural proteins, in such a way as to stimulate cell turnover for the reconstitution of the extracellular matrix and to ensure a long- lasting effect.

[0010] Some known solutions are, for example, described in patent documents EP1064010B1 and US9458197B2, wherein a composition is described comprising a plurality of portions / fragments of elastin chains, typically derived from proteolytic elastin digestion processes including from animal tissue, which simulate elastin activity or stimulate endogenous elastin production.

[0011] In order to meet the need to improve the efficacy of the aforementioned compositions, the Applicant realised that it might be effective to make available not only substances capable of stimulating the endogenous production of the structural proteins of the skin, but also the precursor elements - i.e. the basic constituents - of those proteins, in such a way as to make them directly available in situ and to be able to immediately respond to the stimulus for their endogenous production. Furthermore, the Applicant realised that releasing these precursor elements in a gradual manner would be helpful in prolonging the endogenous production of these structural proteins over time.

[0012] The Applicant has thus found that the combination of polycaprolactone, a recombinant protein of type I collagen and an admixture of amino acid precursors of skin structural proteins, wherein the amino acids are provided in the specific weight ratios of the proteins to be produced, makes it possible to obtain a composition for cosmetic treatment of the dermis that effectively stimulates the endogenous production of these proteins and bio-restructuring of the skin structure, with the aim of stimulating cell turnover and treating skin imperfections.

[0013] Description of the invention

[0014] Therefore, in a first aspect thereof, the present invention is directed to a composition for cosmetic treatment of the dermis. Preferably, the composition comprises polycaprolactone (PCL).

[0015] Preferably, the composition comprises a recombinant protein of type I collagen. Preferably, the composition comprises an admixture of amino acids.

[0016] Preferably, said admixture of amino acids comprises at least one of Glycine, Valine, Proline and Alanine, more preferably it is formed by Glycine, Valine, Proline and Alanine.

[0017] Preferably, the fraction by weight of Glycine with respect to the total weight of the admixture of amino acids is between 25% and 40%.

[0018] Preferably, the fraction by weight of Valine with respect to the total weight of the admixture of amino acids is between 15% and 25%.

[0019] Preferably, the fraction by weight of Proline with respect to the total weight of the admixture of amino acids is between 10% and 20%.

[0020] Preferably, the fraction by weight of Alanine with respect to the total weight of the admixture of amino acids is between 20% and 35%.

[0021] Thanks to these characteristics, elastin-precursor amino acids such as glycine, valine, proline and alanine combine - in appropriate weight ratios - synergistically with the effect of polycaprolactone and recombinant protein of type I collagen, joining together in a composition that effectively stimulates endogenous elastin production. In fact, the Applicant surprisingly found that the above-mentioned recombinant protein not only induces the production of pro-collagen C-peptide type I in fibroblasts, which in turn stimulates the formation of the triple helix of type I collagen within the endoplasmic reticulum, but also stimulates the production of elastin.

[0022] PCL, a biodegradable synthetic polymer derived from the polyesterification of caprolactone acid, induces fibroblasts to neo-collagenesis, i.e. the synthesis of collagen, which is essential for improving skin elasticity and structure. In addition, amino acids provide support for the maintenance of tissue homeostasis and trophism of the subcutaneous environment, promoting the activation of cell renewal processes and regeneration of the extracellular matrix.

[0023] As will be better explained later, polycaprolactone plays a very important role in the composition as it also acts as a barrier for amino acids and recombinant protein of type I collagen against the body's immune reaction, whose phagocytic cells, for instance, attack the composition once injected, recognising it as a foreign body. PCL thus acts as a protective shield for the above- mentioned elements, greatly reducing the patient's inflammatory reaction to the injection of the composition.

[0024] The three above-mentioned components thus cooperate synergistically with one another, making the composition particularly effective in treating signs of skin ageing and loss of volume: in fact, the composition is essentially biorestorative and is suitable for regenerating the body's extracellular matrix. Moreover, the resulting composition appears particularly suitable for injection into a patient's tissues.

[0025] Preferably, the fraction by weight of glycine is between 32% and 36%, more preferably around 34%.

[0026] Preferably, the fraction by weight of valine is between 19% and 23%, more preferably around 21%.

[0027] Preferably, the fraction by weight of proline is between 13% and 18%, more preferably around 15%.

[0028] Preferably, the fraction by weight of alanine is between 27% and 31%, more preferably around 29%.

[0029] The weight ratios of the various amino acids with respect to the total weight of the admixture they form essentially correspond to those with which the same amino acids are found in elastin: in this way, the precursors of this protein are made immediately available, the synthesis of which is stimulated by the interaction of the admixture of amino acids with polycaprolactone and the recombinant protein of type I collagen.

[0030] In some embodiments, the fraction by weight of the admixture of amino acids to the total weight of the composition is between 0.3% and 1.5%.

[0031] Preferably, the fraction by weight of the admixture of amino acids is in the order of 1%.

[0032] Appropriately, similar values of the weight ratios of amino acids and the fraction by weight of their admixture with respect to the composition make it possible to optimise the interaction between the amino acids themselves and the other components (PCL and recombinant protein of type I collagen): in this way, in addition to ensuring effective stimulation of collagen and elastin production, it is possible to avoid remnants of amino acid residues downstream of the interaction between the components.

[0033] In embodiments, the recombinant protein of type I collagen is an R-polypeptide alpha 1 chain, which is advantageously an effective stimulant for type I collagen and elastin synthesis.

[0034] Preferably, the fraction by weight of the recombinant protein of type I collagen relative to the total weight of the composition is between 0.001% and 0.01% of the total weight of the composition. Even more preferably, it is in the order of 0.004%.

[0035] Preferably, the recombinant protein of type I collagen is obtained from transgenic silkworms, e.g. according to the teachings of European Patent EP 1811027.

[0036] In preferred embodiments, the fraction by weight of polycaprolactone in relation to the total weight of the composition is between 3% and 10%, more preferably between 3% and 7%.

[0037] In embodiments, polycaprolactone has an average molecular weight between 4000 Daltons and 6000 Daltons, preferably between 4800 Daltons and 5200 Daltons.

[0038] In preferred examples, polycaprolactone has a substantially spheroidal shape and is preferably in powder form. Preferably, it has an average granulometric size between 20 microns and 70 microns. Even more preferably, the average granulometric size is between 40 and 70 microns.

[0039] These values are determined by laser diffraction, a measurement technology involving a laser beam directed at a group of particles dispersed in a liquid or air stream, and a receiving unit, which detects the light diffraction pattern of the group of particles. The average granulometric size value is defined by the median of the distribution of the granulometric size values of the polycaprolactone particles: in practice, the average granulometric size value is selected in such a way that 50% of the total particles have a granulometric size smaller than this value (or, equivalently, 50% of the total particles have a size larger than it).

[0040] Advantageously, PCL in powder form allows its effective suspension in an injectable matrix in gel form, without the use of solvents. In fact, polycaprolactone in non-spheroidal conformations, e.g. cylindrical or elliptical as in commonly marketed pellets, is not suitable for use in injectable compositions, as it does not guarantee homogeneous distribution within the respective matrix and does not allow extrusion through the appropriate needles / cannulas used for injection, thus compromising the efficacy and usability of the product.

[0041] Polycaprolactone powder has an intrinsic viscosity preferably between 1 dL / 9 and 1.5 dL / g, and more preferably between 1.13 dL / g and 1.38 dL / <3, making it particularly suitable for injectable compositions, as a uniform consistency and easy handling through small needles / cannulas is required. This feature also allows them to meet the requirements of biocompatibility and safety for the body.

[0042] Appropriately, in addition to providing an immediate filling effect on the areas treated by injecting the composition, the PCL it contains has the ability to stimulate the natural production of collagen and elastin, improving the structure and elasticity of the skin.

[0043] In preferred embodiments, the composition has an average viscosity between 24 and 27 Pas, preferably between 25 and 26 Pas, more preferably around 25.5 Pas.

[0044] This advantageously allows the composition to be injected into the body effectively and safely for the patient.

[0045] In some embodiments, the composition comprises 75% to 98%, and preferably 85% to 95%, by weight of at least one carrier which is suitable for the intradermal or subcutaneous administration by injection, in particular at a layer underlying the epidermis, preferably comprising at least partially the hypodermis, i.e. the deepest layer of skin underlying the dermis.

[0046] In preferred embodiments, the composition comprises water. Preferably, the composition comprises a rheological modifier. Preferably, the composition is in gel form.

[0047] Given the insolubility of PCL in water, this characteristic advantageously allows to suspend the undissolved polycaprolactone within a gelled phase, which is used as a carrier to convey the PCL within the composition: in this way, it is possible to avoid the use of solvents, which could be poorly tolerated and significantly increase the risk of bio-incompatibility of the composition, with consequent allergic and inflammatory reactions in the patient's body.

[0048] In some embodiments, the rheological modifier comprises carboxymethyl cellulose (CMC), a cellulose derivative obtained by introducing carboxymethyl groups.

[0049] Appropriately, this characteristic makes CMC water-soluble and gives it thickening, stabilising and film-forming properties.

[0050] Polycaprolactone and carboxymethylcellulose essentially act as a reservoir for the admixture of amino acids and the recombinant protein of type I collagen, as will be better explained later. Together with PCL, CMC also acts as a protective barrier for them, as well as a rheological regulator. Following the introduction of the composition into the body, it is essentially retained in a region close to the injection site. As PCL and CMC are slowly metabolised by the body, fulfilling their function as a physical barrier against metabolic aggression in the patient's body, the admixture of amino acids and recombinant protein of type I collagen are essentially released to stimulate neo-collagenesis, i.e. the endogenous production of skin structural proteins such as collagen and elastin. In this way, gradual consumption of the matrix formed by PCL and CMC ensures slow and gradual release of amino acids and recombinant protein of type I collagen, which are well-tolerated by the patient: this allows, in fact, the metabolisation time to be prolonged (compared to the immediate release of the composition in the patient's body), which considerably reduces the intensity of the inflammatory reaction, and also allows prolonged endogenous stimulation of the skin's structural proteins.

[0051] Preferably, the fraction by weight of carboxymethyl cellulose with respect to the total weight of the composition is between 3% and 10%, and more preferably between 3% and 7%.

[0052] In fact, CMC has gelling properties and is able to form viscous solutions even at low concentrations. Its stabilising properties make it possible to prevent the separation of the components it contains: such a fraction by weight makes it possible to optimise the suspension of PCL within it and to achieve a good level of extrudability of the composition, preventing the formation of an excessively viscous, compact gel that is unsuitable for injection. Moreover, it is a bio- compatible material and is therefore considered safe, non-toxic and well- tolerated by the human body.

[0053] Preferably, the carboxymethylcellulose has a molecular weight between 200 KDa and 300 KDa.

[0054] In preferred embodiments, the weight ratio between carboxymethylcellulose and polycaprolactone is between 0.8: 1 and 1.5: 1. Preferably, their weight ratio is about 1 : 1.

[0055] This allows to obtain optimal mixing, high homogeneity of the compound and, as will be better explained later, effective retention of the properties of the composition downstream of the sterilisation process.

[0056] In preferred embodiments, the composition comprises a buffer agent. Preferably, the buffer agent comprises sodium phosphate dodecahydrate and / or sodium phosphate dihydrate.

[0057] In some embodiments, the composition has a pH between 6 and 8.

[0058] Preferably, the osmolarity of the composition is between 250 and 350 mOsm / L, and more preferably between 280 and 300 mOsm / L.

[0059] Osmolarity and pH are important parameters with regard to product safety, efficacy and performance: they influence tissue response, as well as patient comfort during and after the composition injection procedure. Advantageously, in order to ensure optimal integration in the tissues, the aforementioned osmolarity and pH values are as close as possible to the physiological values of body fluids: in this way, osmotic imbalances that could cause undesirable effects - such as oedemas or tissue dehydration - can be prevented and the risk of inflammation and / or cell damage minimised. In addition, similar osmolarity and pH values help to avoid burning sensations during or after injection. To keep them within optimal parameters, the buffer agent can cooperate, for example, with sodium chloride.

[0060] In embodiments, the composition is suitable for use in the treatment of wrinkles, undulations, laxity, folds and / or furrows by subcutaneous hypodermic injection of a cosmetically active quantity of said composition.

[0061] The term "cosmetically active" means an amount of a composition that is effective and capable of producing visible results in terms of reduction of the aforementioned wrinkles, undulations, laxity, folds and / or furrows compared to a skin condition prior to administration of the composition.

[0062] Injection into the hypodermis makes it possible to introduce the composition into the body by making a small number of entry points, from each of which several administrations can be made. In fact, the deepest layer of the skin is substantially and relatively "empty": using a suitably long injection device, such as a needle-cannula and / or hypodermic needle, it is possible to inject the composition into several regions of the hypodermis from a single hole in the skin, moving the injection device into the subcutaneous layer and releasing the composition in several different areas. This feature advantageously avoids the need for as many micro-injections requiring the creation of as many holes in the skin, thus greatly reducing redness, swelling, inflammation and the generation of unwanted skin imperfections post-injection. It may be sufficient, in fact, to make a small number of injection sites from which to introduce the composition: preferably, the total number of injections made may be four, for example, in the area corresponding essentially to the middle and lower thirds of the patient's face. As mentioned above, the filling of the hypodermis can also provide support for the upper skin layers, which are thereby stretched and invigorated thanks to the endogenous production of collagen and elastin stimulated by the injected composition.

[0063] Preferably, the composition is administered in a single-dose syringe (with a volume of essentially 2.25 ml) filled with between 1.5 ml and 2 ml. More preferably, the desired dosage volume is in the order of 1.8 ml.

[0064] This advantageously makes the composition safe from the point of view of handling and administration, greatly reducing the risk of injection of the wrong dosage volume and accidental contamination of the composition.

[0065] In a second aspect thereof, the present invention concerns the cosmetic use of a composition as described above, for reducing the depth or improving the aesthetic appearance of wrinkles, undulations, laxity, folds and / or furrows, particularly on the face or neck.

[0066] The composition is advantageously achieved by mixing the components mentioned and described so far, which preferably takes place in three separate and successive steps. Specifically, a first step involves mixing the amino acids (glycine, valine, alanine, proline) in the appropriate weight ratios described above with recombinant protein of type I collagen in sterile water for injection. Preferably, the buffer agent and a neutral salt, e.g. sodium chloride, are also added to this aqueous solution: in this way, the values of the pH and osmolarity parameters can be adjusted and brought within the predefined values suitable for the composition of the invention.

[0067] Preferably, the components thus combined are filtered through a membrane filter, e.g. with a pore size of approximately 0.2 microns.

[0068] A second step involves adding the rheological modifier to the filtered solution under vigorous agitation regime, e.g. by means of a high-frequency rotation stirrer, until a substantially homogeneous admixture is obtained.

[0069] Finally, in a third step, polycaprolactone is added to the resulting admixture, maintaining the vigorous stirring regime until a homogeneous final admixture is obtained, e.g. for 10 to 20 minutes, preferably for about 15 minutes. For example, the agitator used may be a mixer with a rotation frequency of approximately 1500 rpm.

[0070] Another critical parameter is also the sterilisation process of the composition, which must be effective not only to ensure its safety for the human body during and post-injection, but also to maintain the integrity of its chemical and physical parameters, even during the subsequent storage step prior to its administration into the subcutaneous tissues.

[0071] Surprisingly, the tests performed showed that the most common sterilisation method - carried out with a hot-wet autoclave cycle - produced effective and non-destructive results with regard to the morphological properties of the composition. Although the melting / softening point of PCL (around 60°) is at a temperature below the maximum temperature reached during sterilisation (around 121°), it was verified that, following the process, the composition retains the integrity of its morphological properties and that the PCL powder remains homogeneously suspended within the carboxymethylcellulose gel matrix. Although a reduction in viscosity was observed post-sterilisation (a phenomenon actually expected and characteristic of heat-treated polymers), the relaxation of the CMC structure resulted in an improvement in its extrudability and flowability, facilitating the introduction of the composition into the tissues and thus ensuring an optimal level of comfort for the patient during the injection step.

[0072] Preferred embodiment

[0073] In a preferred example, the composition for cosmetic treatment of the dermis according to the present invention is shown in Table 1.

[0074] Component c), i.e. the admixture of Glycine, Valine, Alanine and Proline, constitutes 0.79% by weight of the composition. With respect to the total weight of the admixture, the percentages of Glycine, Valine, Alanine and Proline are 34%, 21%, 29% and 15% respectively.

[0075] The composition of this preferred embodiment is obtained by mixing the above- mentioned amino acids, recombinant protein of type I collagen (preferably an R-polypeptide alpha 1 chain), sodium chloride, sodium phosphate dodecahydrate and sodium phosphate dihydrate in sterile water for injection. The components thus combined are filtered through a membrane filter with 0.2 micron pores. Then, by vigorous mixing in a mixer, first the carboxymethylcellulose and then the polycaprolactone are added to obtain a homogeneous final composition ready for sterilisation. The latter is carried out in a hot-wet cycle lasting about 20 minutes, in which the desired sterilisation temperature -about 121°C- is reached after about half the time.

[0076] Table 1

[0077] Experimental results

[0078] In order to test the efficacy of the composition according to the present invention, a plurality of selected patients were treated with the composition of the example described above. In these patients, the composition was administered at areas characterised by a loss of tissue volume, particularly following weight loss. The areas treated for each patient were: the abdominal region, hips, inner thighs and arms.

[0079] For each treated area, an administration zone was identified, at which the dermal composition was injected.

[0080] After a period of 45 to 60 days after the injection of the composition, histological samples were taken from the areas of administration and from respective adjacent untreated areas, resulting in a plurality of so-called "treated samples" and a plurality of respective so-called "virgin samples", respectively.

[0081] The skin tissue of each of the aforesaid samples was divided into three layers: papillary, middle and deep. For each of them, high-resolution scans of the respective histological field were taken, obtaining respective histological images; these images were then analysed by means of an algorithm aimed at identifying the density and organisation of collagen and elastin fibres.

[0082] For each of the histological images of the above-mentioned samples, a plurality of histological parameters descriptive of the analysed tissue were detected and compared with each other.

[0083] The parameters analysed are listed and described below.

[0084] 1. Median angular deviation: it is represented by the angle (measured in degrees) between the axis of extension of the protein fibres and a direction of correct orientation of the fibres with respect to the reference plane defined by the epidermis. For fibres in the middle and deep layers, the correct direction is defined by an axis essentially parallel to the reference plane; for fibres in the papillary layer, the correct direction of orientation is defined by an axis essentially orthogonal to the reference plane. This plane is identified by a plane locally tangent to the tissue of the sample taken, and may for example be coincident with the plane of the relevant histological image. The median angular deviation therefore describes the arrangement of the protein fibres with respect to a relative preferential direction of orientation, depending on the dermal layer to which they belong.

[0085] 2. Percentage of "correctly oriented" fibres: the parameter represents the percentage of protein fibres whose angular deviation from the epidermis reference plane is contained below a predetermined threshold, in this case set at + / - 15° from the plane.

[0086] 3. Coherence index: is a dimensionless parameter expressing the degree of anisotropy of protein fibres. In other words, the coherence index is representative of the degree of mutual alignment between the protein fibres of the analysed portion of the tissue.

[0087] 4. Fragmentation index: is determined by evaluating the length of the protein fibres and the number of endpoints per megapixel of the analysed histological image. In practice, the fragmentation index is a measure of fibre length.

[0088] The results of the analysis conducted are represented by the graphs illustrated in the accompanying Figures, in which:

[0089] - Figure 1 represents the median angular deviation of protein fibres found in treated and virgin samples,

[0090] - Figure 2 represents the percentage of correctly oriented protein fibres in treated and virgin samples,

[0091] - Figure 3 represents the distribution of the protein fibre coherence index in treated and virgin samples, and

[0092] - Figure 4 represents the protein fibre fragmentation index in treated and virgin samples.

[0093] The results presented in the graphs in Figures 1 to 4 are in the form of a box- and-whisker plot, or in short, a boxplot. In each graph, the y-axis shows the parameter analysed and comprises:

[0094] - a first boxplot, labelled "PRE", representing data on virgin samples, untreated with the composition of the invention;

[0095] - a second boxplot, labelled "POST", representing the data on the treated samples, analysed between 45 and 60 days after administration of the composition of the invention to the patient.

[0096] Each boxplot comprises a rectangle (the "box") delimited at the bottom by the first quartile and at the top by the third quartile of the data distribution under analysis. More specifically, the first and third quartiles represent the values within which 25% and 75% of the analysed data fall, respectively. The width of the box along the y-axis is equal to the inter-quartile range, i.e. the difference between the third and first quartile; moreover, within it, the box is divided into two regions by the median, a value within which 50% of the data falls.

[0097] Two segments (the "whiskers") extend vertically from the rectangle of each boxplot, delimited respectively by the minimum and maximum values of the distribution, from which anomalous values, defined as outliers, have previously been excluded. Outliers are values that deviate significantly from the rest of the data in the distribution and are represented graphically as dots: in particular, they were identified by setting a threshold value, from the first and third quartiles, corresponding to 1.5 times the interquartile range.

[0098] With particular reference to Figure 1, the y-axis shows the median angular deviation of the protein fibres of the analysed tissues with respect to their correct direction of orientation. In practice, for each histological image acquired, the median value of the angle between the axis of the fibres of the skin tissue analysed and the reference plane of the epidermis or an axis orthogonal to it was determined, depending on whether the fibres analysed belonged to the medium-deep or papillary dermal layer.

[0099] In order to achieve a compact and uniform skin appearance, it is crucial that the protein fibres have an orientation as parallel as possible to their correct direction of orientation, so as to ensure a more even distribution of mechanical stresses and the maintenance of a smoother and more regular surface. On the contrary, irregularly arranged fibres can lead to inhomogeneity in the dermal structure, promoting the formation of micro-wrinkles. Therefore, it is desirable that the orientation of the fibres is as close as possible to the correct orientation direction. As can be seen from the graph in Figure 1, the distribution of the average angular deviation of the protein fibres post-treatment has a smaller median value than the median value of the angular deviation of the corresponding virgin tissue, in which a significant percentage of fibres are irregularly and incorrectly oriented with respect to the reference direction. Thus, it can be seen that, downstream of the administration of the composition, the distribution of the protein fibres in the respective layers is more regular overall : in particular, the distribution in the middle and deep layers is more organised and more concordant with the plane of the epidermis, while the distribution of the fibres in the papillary layer is more concordant with an axis orthogonal to the plane itself, resulting in a smoother and more compact aesthetic skin appearance.

[0100] Figure 2 shows a comparison between the percentage of correctly oriented fibres in the treated and virgin samples. As anticipated, the threshold value for the angular deviation of the fibres was selected as + / - 15° from the reference plane of the epidermis. In agreement with the results illustrated in Figure 1, it can be seen that the percentage of correctly oriented fibres is higher downstream of the treatment, as the median value of the angular deviation is overall smaller and the fibres are overall more organised and concordant with the reference plane of the epidermis.

[0101] Figure 3 illustrates the distribution of the coherence index, which represents the mutual organisation and arrangement of protein fibres in the tissues of treated and virgin samples. A bundle of fibres oriented anisotropically, i.e. aligned along a prevailing direction, gives the dermal tissue a more regular surface appearance and greater support to the epidermis, reducing the formation of micro-folds, hollows and undulations.

[0102] As depicted in the graph, the distribution of the post-treatment coherence index has a higher median value than the median value obtained when analysing the virgin samples. A high coherence index represents an organised extracellular matrix in which the protein fibres are arranged in a network with directional continuity. It is therefore clear that administering the composition allows the fibres to be reorganised by arranging them in a more orderly manner than in the pre-treatment condition. In addition, the composition promotes effective tissue regeneration: more organised distribution of protein fibres guides the orientation of new fibres produced by fibroblasts, facilitating the orderly reconstruction of dermal tissue, reducing the risk of inhomogeneous fibrous networks and resulting in a more effective biological scaffold.

[0103] With particular reference to the graph in Figure 4, the boxplots illustrating the distribution of pre-treatment and post-treatment data are representative of the fragmentation of protein fibres in the tissues of the samples analysed. The y- axis shows the number of endpoints per megapixel (Mpx), i.e. the number of fibre endpoints detected in a predetermined area of the analysed histological image. The greater the number of endpoints detected, the greater the degree of fragmentation: short, disorganised fibres compromise the continuity and density of the extracellular matrix and promote the appearance of surface irregularities. In order to achieve a good level of skin firmness, it is therefore desirable that the fibres are long and orderly: a low level of fragmentation, in fact, gives the tissue a more continuous and coherent structure, which reduces sagging points and is able to support the epidermis evenly. The length of the fibres is also related to the tissue's ability to withstand external stresses; in fact, a network of longer fibres opposes tissue deformation more effectively than shorter segments of fragmented fibres.

[0104] Long, continuous fibres also contribute to the creation of a smooth and homogeneous skin profile, preventing the appearance of micro-depressions, unevenness and loss of support of the epidermis substantially at the points where the fibres are interrupted.

[0105] Figure 4 shows how the median value of the protein fibre endpoints is significantly reduced in the post-treatment condition compared to the pretreatment condition: samples treated with the composition according to the invention exhibit in fact an extremely reduced degree of fragmentation compared to the corresponding virgin samples and thus have longer and more continuous protein fibres.

[0106] It should be noted that the effect of the injectable composition described here is essentially a long-term regeneration of the extracellular matrix.

[0107] Specifically, the Applicant observed that substantially in the first week after administration, there is an immediate response due to the hydrating component and initial tissue stimulation, consisting of improved skin hydration, as well as a noticeable increase in skin turgidity and a fuller, firmer texture.

[0108] Substantially, about a month after administration, a primary biological response of the fibroblasts to the injected stimulant substance occurs, and they are activated by producing new protein fibres and reorganising the extracellular matrix, thus initiating an initial neo-collagenesis step that leads to an increase in skin elasticity and initial smoothing of fine wrinkles.

[0109] Substantially about three months after administration, the biostimulation step starts and the treatment shows more visible structural effects: the production of new collagen and elastin fibres promotes partial restoration of volume in areas with poor tonicity, as well as a reduction of deeper undulations and an improvement in the definition of specific areas, such as the jawbone and / or cheekbones.

[0110] Substantially between 3 and 24 months after administration, tissue remodelling is consolidated and stabilisation of the extracellular matrix structure occurs, with an increase in dermal density and skin tone without significant migration of composition or swelling. In fact, the substance is not volumising in nature, but bio-restructuring, and the actual regeneration of the extracellular matrix is completed during this time. A progressive consumption of the matrix formed by PCL and CMC, in fact, ensures slow and gradual release of amino acids and recombinant protein of type I collagen, prolonging the metabolisation time and extending the endogenous stimulation of the skin's structural proteins.

[0111] In view of the foregoing, the Applicant has verified that the composition for cosmetic treatment of the dermis according to the present invention achieves effective stimulation of endogenous elastin and collagen production by synergistic interaction of the recombinant protein of type I collagen with elastin precursor amino acids and with polycaprolactone. It is also verified that the resulting composition, injected into the hypodermis, allows to create structural support for the upper skin layers, relaxing the skin and making it firmer and more toned. Moreover, the bio-revitalising action of the composition not only makes the effect last longer, thanks to the natural production of the skin's structural proteins and cell turnover processes, but also looks more natural.

Claims

Claims1. A composition for cosmetic treatment of the dermis comprising: a) polycaprolactone, b) b) a recombinant protein of type I collagen and c) c) an admixture of amino acids consisting of glycine, valine, proline and alanine, wherein, with respect to the total weight of the admixture of amino acids, the fraction by weight of glycine is between 25% and 40%, the fraction by weight of valine is between 15% and 25%, the fraction by weight of proline is between 10% and 20% and the fraction by weight of alanine is between 20% and 35%.

2. A composition according to claim 1, wherein, with respect to the total weight of the admixture of amino acids, the fraction by weight of glycine is between 32% and 36%, the fraction by weight of valine is between 19% and 23%, the fraction by weight of proline is between 13% and 18% and the fraction by weight of alanine is between 27% and 31%.

3. A composition according to claim 1 or 2, wherein the fraction by weight of the admixture of amino acids with respect to the total weight of the composition is between 0.3% and 1.5%.

4. A composition according to any one of the preceding claims, wherein the recombinant protein of type I collagen is an R-polypeptide alpha 1 chain.

5. A composition according to any one of the preceding claims, wherein the fraction by weight of the recombinant protein of type I collagen with respect to the total weight of the composition is between 0.001% and 0.01% of the total weight of the composition.

6. A composition according to any one of the preceding claims, wherein the fraction by weight of the polycaprolactone with respect to the total weight of the composition is between 3% and 10%, preferably it is between 3% and 7%.

7. A composition according to any one of the preceding claims, wherein the polycaprolactone has an average molecular weight between 4000 Dalton and 6000 Dalton, preferably between 4800 Dalton and 5200 Dalton.

8. A composition according to any one of the preceding claims, wherein the polycaprolactone is in the form of powder having an average granulometricsize between 20 micrometres and 70 micrometres, preferably between 40 and 70 micrometres, wherein said average granulometric size is the median of the granulometric size values distribution of said polycaprolactone powder, said values being determined by laser diffraction.

9. A composition according to any one of the preceding claims, having an average viscosity between 24 and 27 Pas, preferably between 25 and 26 Pas.

10. A composition according to any one of the preceding claims, comprising from 75% to 98% by weight of at least one carrier which is suitable for the intradermal or subcutaneous administration by injection.

11. A composition according to any one of the preceding claims, comprising water and a rheological modifier.

12. A composition according to the preceding claim, wherein the rheological modifier comprises carboxymethyl cellulose.

13. A composition according to the preceding claim, wherein the fraction by weight of the carboxymethyl cellulose with respect to the total weight of the composition is between 3% and 10%, preferably between 3% and 7%.

14. A composition according to claim 12 or 13, wherein the weight ratio between the carboxymethyl cellulose and the polycaprolactone is between 0.8: 1 and 1.5: 1, preferably it is approximately 1 : 1.

15. A composition according to one or more of claims 12 to 14, wherein the carboxymethyl cellulose has a molecular weight between 200 KDa and 300 KDa.

16. A composition according to any one of the preceding claims, comprising a buffer agent.

17. A composition according to the preceding claim, wherein the buffer agent comprises sodium phosphate dodecahydrate and / or sodium phosphate di hydrate.

18. A composition according to any one of the preceding claims, having a pH between 6 and 8.

19. A composition according to any one of the preceding claims, having osmolarity between 250 and 350 mOsm / L, preferably between 280 and 300 mOsm / L.

0. Use of a composition according to any one of the preceding claims for the treatment of wrinkles, undulations, laxity, folds, furrows by subcutaneous hypodermic injection of the composition.

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