• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    VISCOELASTIC GELS AS INNOVATIVE LOADS. The present invention relates to biomaterials obtainable by mixing: - the auto-reticulated derivative of acihi (ACP) with - the derivative (HBC) of hyaluronic acid cross-linked with - 1,4-butanediol diglycidyl ether (BDDE) in the weight ratio between 10:90 and 90:10, as new fillers.
    公开号:BR112012004157B1
    申请号:R112012004157-6
    申请日:2010-08-25
    公开日:2021-03-09
    发明作者:Matteo D'este；Davide Renier
    申请人:Fidia Farmaceutici S.P.A.；
    IPC主号:
    专利说明:

    FIELD OF THE INVENTION
    [001] Hyaluronic acid (HA) is a hetero-polysaccharide consisting of alternating residues of D-glucuronic acid and N-acetyl-d-glucosamine.
    [002] HA is a straight chain polymer with a molecular weight ranging between 50,000 and 13 x 106 Da, depending on the source from which it comes and the preparation methods employed.
    [003] HA is present in nature in pericellular gels, in the fundamental substance of the vertebrate connective tissue (of which it is one of the main components), in the vitreous humor and in the umbilical cord.
    [004] HA plays an important role in the biological organism as a structural and mechanical support for tissues, and as an active component in the cellular physiology of tissues such as skin, tendons, muscles and cartilage, being one of the main molecules in the matrix cartilaginous, also representing the main non-protein constituent of synovial fluid. As it is a highly hydrophilic viscoelastic molecule, it provides lubricating properties to the synovial fluid. HA has therefore been used in osteoarthritis for about 30 years, mainly to treat associated pain.
    [005] Hyaluronic acid (HA) still plays a crucial role in the tissue repair process from a structural point of view (in the organization of the extracellular matrix and regulation of its hydration), and as a regulatory / stimulating substance in a wide range of physiological processes in which said polysaccharide acts directly and / or indirectly (clot formation, phagocytic activity, fibroblast proliferation, neovascularization, re-epithelialization, etc.) (Weigel P. et al., J Theoretical Biol, 1986: 219 -234; Abatangelo G. et al., J Surg Res, 1983, 35: 410-416; Goa K. et al., Drugs, 1994, 47: 536-566). As these properties have long been recognized, HA is also used for the preparation of bandages in the care of wounds, ulcers and skin lesions of various origins.
    [006] Hyaluronic acid is also used as a filler for wrinkles, creases and small depressions of the face, and also to increase the volume of the lips and face, due to being immunologically inert, non-toxic, biodegradable and bioresorbable.
    [007] Treatment based on hyaluronic acid is indicated for the correction of: volume and contour of the lips; creases (for example nasolabial folds; remodeling of facial contours (for example, face and chin); wrinkles (for example, glabellar lines and oral commissures); periorbital wrinkles; post-acne fibrous scars; post-traumatic fibrous scars; tissue defects soft, rhinoplasty scars.
    [008] Hyaluronic acid is not a permanent filler, meaning that once injected, the product is gradually metabolized and reabsorbed by the body, over time, varying according to the area treated and the type of preparation used. The filling and swelling effect (or wrinkle attenuation) is immediate, lasting only a few weeks. The main products on the market can be classified under the following categories, based on their different resorption times: quick resorption fillers (2 to 3 months), medium term resorption fillers (5 to 6 months), resorption fillers slow (1 year) as Restylane Sub Q (QMed, EP0839159).
    [009] In the dermis, HA performs moisturizing functions due to its great ability to bind to water and structural functions such as "scaffolding", because, by the connection with other substances, it forms macromolecular complexes that make the skin compact.
    [010] The mechanism of action, therefore, consists of immediate volumetric filling, due to the product's viscoelastic properties and new collagen synthesis, due to the stimulation of cutaneous fibroblasts.
    [011] However, HA is a natural polysaccharide that is quickly broken down by hyaluronidase enzymes present in connective tissue; in order to obtain fillers whose effect lasts for several months, HA is therefore subjected to crosslinking processes that improve its viscoelastic properties and increase its residence time. The fillings thus formed are cross-linked, for example, by means of BDDE (1,4-butanediol diglycidyl ether, Restylane®, BELOTERO® and Regenyal Idea) or DVS (divinyl sulfone, Hylaform®), which creates bridges between the molecules polymer. However, with the increase in the degree of crosslinking, there is a progressive denaturation of HA to the point of profoundly modifying its physical, chemical and biological properties. Excessively cross-linked HA matrices present as solid particles that are no longer recognized by cells (and especially by the immune system) as HA; the polysaccharide is therefore perceived as a foreign body, which triggers inflammatory reactions with the formation of fibrous capsules around it. Furthermore, excessively cross-linked HA is unable to stimulate the regeneration of dermal / skin tissue, induced as is known from well-established scientific results by HA fragments (especially those with a low molecular weight) that have the effect of stimulating the synthesis of collagen by cutaneous fibroblasts.
    [012] Fills are further classified as resorbable or permanent. Resorbable fillers are the most biocompatible; they consist of hyaluronic acid or collagen, whether modified or present in the native form, and are consequently reabsorbed within a year at most. The permanent type consists of synthetic polymers such as polyacrylamides, particular cross-linked molecules, which form a stable gel when combined with water. The permanent type always remains in situ and is very useful for filling the lips, but its use is not recommended, because the occurrence of increasingly frequent acute inflammations, due to its cutaneous insertion, leads to the formation of fibrous capsules around the filling, what is perceived as a foreign body, which is why it is considered toxic.
    [013] The depositor has perfected a new type of biomaterial as an innovative filler, and / or as a new product for body shaping, formed by mixing two cross-linked HA derivatives in different, yet complementary ways, to obtain a skin substitute / tissue that allows immediate hydration (and consequently, immediate filling) of the treated tissue / skin (a), while at the same time maintaining very long break times in vivo in order to eliminate the need for repeated injections, thus reducing the effects collateral.
    [014] The new biomaterials to which the present invention refers, have particular innovative characteristics of biocompatibility identical to those of hyaluronic acid as such, but their biodegradability is different; when implanted in vivo, its residence time is much longer than that of unmodified HA, thus allowing immediate regeneration / reconstruction of dermal / skin tissue that has lost its original density. DETAILED DESCRIPTION OF THE INVENTION
    [015] The depositor has perfected a new type of biomaterial as a new filler and / or as a new product for body molding, based on the mixture of HA derivatives with different, but complementary, characteristics in obtaining a new product for injection, in the treatment of skin imperfections, in dermatology, in dermocosmetology and / or in cosmetic surgery, producing: immediate dermocutaneous hydration; Immediate filling of the treated tissue; very long break times in vivo; reduced side effects.
    [016] The new biomaterials consist of:
    [017] Self-crosslinking hyaluronic acid (ACP) or HA hexadecylamine (HYADD) mixed with BDDE crosslinked hyaluronic acid (HBC).
    [018] The ACP used in the present invention, prepared as described in EP 0341745, has an average degree of crosslinking between 4 and 5% and is preferably prepared using HA with an average molecular weight (MW) of 200 KDa. When hydrated it presents itself as a self-crosslinking gel without molecules foreign to the native polysaccharide, because it arises from the ester bond between the carboxyl and hydroxyl groups of the same polysaccharide chain and / or adjacent chains. It is, therefore, devoid of immunotoxicity, as biocompatible with native HA, highly humectant, and easily degradable by hyaluronidases, releasing molecules with a low molecular weight, capable of stimulating the synthesis of collagen to improve the tone and elasticity of the skin tissue.
    [019] HA hexadecylamine (HYADD) is prepared as described in EP 1095064 and EP1853279, preferably using HA with an average molecular weight (MW) of 500-730 KDa. with an average degree of final amidation / substitution between 1 and 3 mol%.
    [020] ACP and HYADD are the HA derivatives responsible for immediate hydration (leading to instantaneous dermal filling) proposed by the intradermal injection of the filler to which the present invention refers.
    [021] HA crosslinked with BDDE (a molecule containing epoxy groups to form ethers in the primary hydroxyls of HA) contains the crosslinking molecule and is therefore more resistant to enzymatic degradation, because it has ether bridges that stabilize the polysaccharide giving the product obtained a long residence time.
    [022] Mixing the two types of crosslinked HA results in the formation of a new biomaterial that has biocompatibility characteristics identical to those of native hyaluronic acid, but with a different biodegradability, so that, when implanted in vivo, its time of residence is much greater than that of unmodified HA, thus allowing the regeneration / reconstruction of dermal tissue that has lost its original density. The Depositor further demonstrated that its very unexpected association leads to a much longer breakdown time in vivo than that of commercial reference fillers formed by the same type of HA crosslinked with BDDE, with a consequent increase in residence time. Finally, the Depositor claims the use of new biomaterials as fillers and / or as new products for body shaping in the treatment of skin imperfections, in dermatology, dermocosmetology and / or in cosmetic surgery.
    [023] The chemically heterogeneous nature of the new biomaterials allows the properties of the finished product to be modulated by adequate variation in the weight ratio between the constituents. The two HAs can be mixed in ACP (or HYADD ratio): HBC from 10:90 to 90:10: The weight ratio will be selected on the basis of the desired final viscosity, depending on the treated site. If some implantation areas that need large amounts of biomaterial are going to be treated, as in cases of filling the breasts, buttocks, face or chin, or deep expression wrinkles, the biomaterial used will consist, preferably of good density, and therefore a viscosity adequate in obtaining a gel with excellent consistency and low biodegradability speed; in this case the ACP (HYADD: HBC mixture will be between 10:90 and 50:50, and preferably, 25:75, because the product obtained by increasing the weight fraction of HBC is more suitable to achieve a long-term volume-intensifying effect. duration.
    [024] However, if the treatment is for lip creases or fine wrinkles on the forehead, the ACP (or HYADD): HBC ratio will preferably be between 90:10 and 50:50, since a greater fraction of ACP in the filling produces a material more suitable for the biorevitalization of the skin and correction of fine lines, secondary expression wrinkles and others. In addition, the needle must have a large caliber, the gel must therefore be easily extrudable and less viscous than described above. The rheological properties of the product are therefore adjustable based on the selected ACP: HBC ratio.
    [025] ACP (or composition HYADD) / HBC being the same, the properties of the biomaterial can also be adequately modulated by means of a target selection of the vehicle, in which it is prepared: for example, a mixture by weight of ACP: HBC 50:50 dispersed in saline (0.9% NaCl) will be more viscous than if dispersed in phosphate buffer at pH = 6.95; and therefore, for this specific mixture, saline is a more suitable medium for formulating products with a limited rate of dispersion in situ.
    [026] Materials consisting mainly of HBC will exhibit the opposite profile. The viscoelastic properties of the material consequently affect the product's performance.
    [027] The present invention also relates to processes for preparing the two biomaterials described above: process A and process B.
    [028] The innovative processes A and B are divided into two stages: 1. process for producing the HBC derivative, and 2. process for mixing it with the ACP or HYADD derivative.
    [029] The two stages lead to the production of products with a high degree of purity; With the methods normally employed in the production of crosslinked HA with BDDE, the purification processes are carried out by washing the obtained gel mass, or by dialysis. In both cases, the optimal purification efficiency cannot be achieved, due to the nature of the gel matrix, which, due to its tendency to swell, incorporates large amounts of solvent. These gels have low mobility and transport capacity, tending to precipitate like gelatinous gums. The precipitate thus obtained, isolated as a solid, has different properties of solubility and rheology when rehydrated, especially the ability to swell, elasticity and homogeneity (characteristics essential for filling) of the gel before purification.
    [030] However, the method described below by the Depositor as process A precipitates the product in the form of a finely divided powder, which is then easily washable. In addition, the careful choice of reaction conditions produces, after isolation by precipitation and washing, a product capable of reconstructing the gel, through rehydration and sterilization, resulting in a biomaterial with reproducible characteristics, well standardized in terms of elasticity and homogeneity.
    [031] Process B does not include the precipitation step of the HBC product as a powder; the purification and homogenization of the gel (obtained after mixing with HBC with ACP or HYADD) is carried out in the crushing step involving the passage of it through a filter with a coefficient of retention of particulate matter between 25 and 150 μm. This step purifies the final gel making it perfectly homogeneous.
    [032] The HA used in the present invention for the preparation of the derivatives described above (HBC, ACP and HYADD) can be derived from any source, such as cock-crest (botanical) extraction or fermentation having an average molecular weight between 400 and 3x 106 Da, preferably between 1x 105 Da and 1x 106 Da, and even more preferably between 200,000 and 1x 106 Da.
    [033] A new manufacturing process A comprises the following steps: Synthesis of crosslinked HBC 1. Dissolution in alkaline solution (preferably 0.15M - 0.35M NaOH) of BDDE diepoxide in a stoichiometric ratio between 2.5 and 25% molar, preferably between 5 and 15 mol% (depending on the intended use of the product; the higher the percentage of BDDE, the longer the residence time) of the repetitive units of hyaluronic acid, followed by: 2. dispersion of HA in the solution referred to in preceding paragraph at room temperature. The HA concentration should be between 80 and 300 mg / ml and the homogenization time between 30 and 300 minutes. 3. Performing the reaction by thermal activation, the solution is heated to a temperature between 35 and 55 ° C between 2 and 36 hours. 4. Extrusion of the mass obtained through a metal sieve, in order to reduce it to particles with a size of approximately 600 μm. 5. Hydration of the gel by diluting it with water by a factor of 3 to 25, for a time between 4 and 48 hours at a temperature of 4 to 24 ° C. 6. Correction of the pH to neutral, with an aqueous solution of HCl having a concentration of 0.5 to 5 moles / L, preferably 1 to 2 moles / L. 7. Addition of 2.5 volumes of water-soluble organic solvent such as ethanol, methanol, isopropanol, n-propanol, dioxane, acetonitrile, acetone and / or mixtures thereof (preferably ethanol and acetone), until the product is obtained in the form of a powder precipitate. 8. Washing with organic solvents such as ethanol, methanol, isopropanol, n-propanol, dioxane, acetonitrile, acetone and / or mixtures thereof (preferably ethanol and acetone) containing an aqueous fraction below 35%. 9. Vacuum drying at a temperature between 30 and 45 ° C for 2 to 7 days, and in any case, until residual solvents are eliminated under 400 ppm to obtain a white HBC powder. Mixing of ACP (or HYADD) with HBC 10. Mixing of HBC powder with ACP (or HYADD) powder in an ACP: HBC ratio between 10:90 and 90:10 (depending on the chosen use, as previously described). 11. Hydration with saline or phosphate buffer, preferably saline (which may contain more excipients such as lidocaine), leading to a total concentration of HA between 12 and 27 mg / ml, preferably between 20 and 25 mg / ml, at a temperature between 0 and 26 ° C. 12. Extrusion through a sieve with a mesh between 50 and 500 μm, preferably between 100 and 250 μm. Filtration is carried out at room temperature, or at a temperature between 25 and 65 ° C, preferably between 40 and 60 ° C. 13. Filling the syringes, preferably made of glass or polymeric material, with the product obtained. 14. Thermal sterilization with saturated steam at a temperature between 120 and 124 ° C (preferably 121.5 ± 1 ° C) for at least 10 minutes.
    [034] New Manufacturing Process B comprises the following steps: Synthesis of crosslinked HBC 1. Dissolution in alkaline solution (preferably 0.15M - 0.35M NaOH) of BDDE diepoxide in a stoichiometric ratio of 2.5 to 25 mol%, preferably between 5 and 15 mol% depending on the intended use of the product) of the repetitive units of hyaluronic acid followed by: 2. Dispersion of HA in the solution referred to in the preceding paragraph at room temperature. The HA concentration should be between 80 and 300 mg / ml and the homogenization time between 30 and 300 minutes. 3. Performing the reaction by thermal activation, the solution is heated to a temperature between 35 and 55 ° C between 2 and 36 hours. 4. Correction of PH to neutral with an aqueous solution of HCl having a concentration of 0.05 to 1 mol / l, preferably 0.1 mol / l. 5. Hydration of the gel by diluting it with water by a factor of 3 to 20 for a time between 4 and 48 hours at a temperature of 4 to 24 ° C. This solution may contain other excipients such as NaCl, sodium or potassium salts of phosphoric acid and lidocaine, preferably in the form of the hydrochloride salt. The sodium salts (chlorine or phosphate) have the function of maintaining the adequate osmolarity of the product, maintaining the pH at a value compatible with the tissues. In a preferred embodiment, from the invention NaCl is added in such a proportion that the final solution contains a concentration between 0.8 and 1.0%, preferably 0.9%; lidocaine hydrochloride, if present, is added in such a proportion that the final formulation contains a proportion between 2.2 and 3.2 mg / ml thereof, preferably 2.7 mg / ml. Mixing ACP (or HYADD) with HBC 6. Mixing HBC gel with ACP powder (or HYADD) in an ACP (or HYADD) ratio: HBC between 10:90 and 90:10 (by weight of the active ingredient) depending on intended use for the new filling, as previously described. Alternatively, ACP or HYADD can be mixed with HBC starting with both bottles in gel form, using a standard stirring system (preferably with an orbital blade) for a time between 30 minutes and 24 hours at a temperature between 0 ° C. and 26 ° C. 7. Crushing and homogenization passing through a filter with a retention coefficient of particulate material between 25 and 150 μm, preferably between 40 and 110 μm. If the viscosity is excessive, the operation can be carried out hot, at a temperature between 25 and 65 ° C. 8. Filling the syringes, glass or polymeric material, with the product obtained. 9. Hot sterilization of saturated steam at a temperature between 120 and 124 ° C (Preferably 121.5 ± 1 ° C) for at least 10 minutes.
    [035] Some examples of preparation in the new filling according to the invention are described below, by way of example and not by way of limitation. Example 1: Synthesis of HBC 500 (HA 500-730 kDa) process A
    [036] 0.075 mol of HA with a molecular weight of 500-730 kDa, produced by fermentation, are dispersed in 215 ml of a 0.25 M NaOH solution containing 1.41 ml of BDDE. The mixture is then heated to 42 ° C and reacted for 3 hours, after which it is hydrated for 24 h with 300 ml of a solution containing a stoichiometric amount of HCl in order to adjust the pH to neutral pH. The total volume is made up of 750 ml and precipitated with 2.5 volumes of ethanol to obtain a settable, filterable precipitate. The mixture is washed with 75% ethanol until exhaustive purification, verified by measuring the specific conductivity of the washing solvents, which must be below 30 μS / cm, being dried under vacuum at 40 ° C for 5 days. The HBC 500 product is obtained with a 87% weight yield. Example 2: Synthesis of HBC 1000 (HA 1MDa) process A
    [037] 1.60 g of HA with an average molecular weight of 1 MDa, produced by fermentation is dispersed in 20 ml of a 0.25M NaOH solution containing 75 μl of BDDE. The mixture is then heated to 42 ° C and reacted for 2 hours. The mixture is hydrated for 24 hours with 20 ml of a solution containing a stoichiometric amount of HCl to adjust the pH to neutral pH. The total volume is made up of 75 ml and the HBC is precipitated with 2.5 volumes of ethanol to obtain a settable, filterable precipitate. The mix is washed with 75% ethanol until exhaustive purification, which was verified by measuring the specific conductivity of the washing solvents, which should be below 30 μS / cm, being dried under vacuum at 40 ° C for 5 days. HBC 1000 product is obtained with a 90% weight yield. Example 3: Synthesis of HBC 200 (HA 200 kDa) process A
    [038] 2.55 g of HA with an average molecular weight of 200 KDa, produced by fermentation, are dispersed in 20 ml of a 0.25M NaOH solution containing 63 μl of BDDE. The mixture is then heated to 42 ° C and reacted for 150 mit. The mixture is then hydrated for 24 hours with 20 ml of a solution containing a stoichiometric amount of HCl. The total volume is made up of 75 ml and precipitated with 2.5 volumes of ethanol to obtain a settable, filterable precipitate. The mixture is washed with 75% ethanol until exhaustive purification, which was verified by measuring the specific conductivity of the washing solvents, which should be below 30 μS / cm, being dried under vacuum at 40 ° C for 5 days. The HBC 200 product is obtained with a 85% weight yield. Example 4: Preparation of ACP gel: HBC 500 in the ratio of 50:50 process A
    [039] 1.00 g of HBC 500, prepared as described in example 1, is mixed with 1.00 g of HA ACP internal ester. The powder is hydrated with 100 ml of 0.9% sterile weight / volume saline at a temperature of 8 ° C for 16 hours. The obtained gel is heated to 48 ° C and filtered through a metal sieve with a 0.17 mm mesh, and then distributed between 1 ml glass syringes, which then pass through a sterilization cycle with saturated steam at a temperature of 121 ° C for 10 minutes. A homogeneous sterile gel suitable for local administration is obtained. Example 5: preparation of ACP gel: HBC 1000, in the ratio of 30:70 process A
    [040] 1.40 g of HBC 1000, prepared in example 2, is mixed with 0.60 g of HA ACP internal ester. The powder is hydrated with 100 ml of sterile 0.9% w / v saline at a temperature of 8 ° C for 16 hours. The obtained gel is heated to 48 ° C and filtered through a metal sieve with a 0.17 mm mesh, and then distributed between 1 ml glass syringes, which pass through a sterilization cycle with saturated steam at a temperature of 121 ° C for 10 minutes. A homogeneous sterile gel is obtained for local administration. Example 6: preparation of ACP gel: HBC 500, in the ratio of 25:75 process A
    [041] 1,875 g of HBC 500, prepared in example 1 is mixed with 0.625 g of ACP of internal HA ester. The powder is hydrated with 100 ml of sterile 0.9% w / v saline at a temperature of 8 ° C for 16 hours. The obtained gel is heated to 48 ° C and filtered through a metal sieve with a mesh size of 0.19 mm, and then distributed between 1 mm glass syringes, with subsequent sterilization cycle with saturated steam at a temperature of 121 °. C for 12 minutes, obtaining a homogeneous sterile gel suitable for local administration. Example 7: preparation of ACP gel: HBC 1000, in the ratio of 75:25 process A
    [042] 0.50 g of HBC 1000, prepared as described in example 2, is mixed with 1.50 g of internal HA ester ACP. The powder is hydrated with 100 ml of sterile 0.9% w / v saline at a temperature of 8 ° C for 24 hours. The obtained gel is heated to 42 ° C and filtered through a metal sieve with a mesh size of 0.17 mm, and then distributed between 2 ml glass syringes with subsequent sterilization cycle with saturated steam at a temperature of 121 ° C for 12 minutes, obtaining a homogeneous sterile gel, suitable for local administration. Example 8: preparation of gel HYADD: HBC 500, in the ratio of 60:40 process A
    [043] 1.20 g of HBC 500, prepared as described in example 1, is mixed with 0.80 g of HA hexadecylamide (HYADD). The powder is hydrated with 100 ml of sterile 0.9% w / v saline at a temperature of 8 ° C for 24 hours. The obtained gel is heated to 52 ° C and filtered through a metal sieve with a mesh size of 0.17 mm, being then distributed between 1 ml glass syringes with subsequent sterilization cycle with saturated steam at a temperature of 121 ° C for 11 minutes, obtaining a homogeneous sterile gel, suitable for local administration. Example 9: preparation of HYADD: HBC 500 gel, in the ratio 40:60 process A
    [044] 8.0 g of HA sodium salt with an average molecular weight of 500-730 kDa, produced by fermentation are dispersed in 40 ml of a 0.25 M NaOH solution containing 0.44 ml of BDDE. The mixture is heated to 41.5 ° C for 2 hours and 40 minutes, after which it is hydrated overnight with 100 ml of a 0.1 M HCl solution and 200 ml of water. 50 ml of a saturated NaCl solution is added and the mixture is allowed to swell overnight. On the next day, 170 ml of acetone and 30 ml of saturated NaCl solution are added, the mixture being precipitated by slowly adding 1 liter of ethanol. The precipitate is washed with the same solvent until the NaCl residues are eliminated, then dried in the oven at 35 ° C under vacuum until the residual solvents are eliminated. The HBC powder thus obtained is mixed in the ratio of 5: 3 with HYADD, prepared as described in patent EP1853279. The mixed powders are hydrated with saline, making a total concentration of 20 mg / ml (corresponding to 12.5 mg / ml of HBC and 7.5 mg / ml of HYADD4). The product is allowed to swell overnight at 5 ° C, and the next day it is filtered through a flat membrane with a nominal particulate matter retention rate of 100 μm. Glass syringes of 1 l are filled with the product thus obtained and sterilized in a cycle with F0 = 13 at 121.5 ° C. Example 10: Skin filling and tolerability of HYADD: HBC gel in the model of intradermal administration to rabbits
    [045] The purpose of the experiment was to evaluate the skin filling, the rush of any macroscopic adverse events and the tissue response induced by the HYADD: HBC gel (prepared as described in example 9) injected into rabbit intradermal tissue, by comparison with the commercial BELOTERO® filler.
    [046] For the said assessment, the tested gels were administered intradermally to male NZW-KBL rabbits weighing 1.8-2.3 kg. Experiment Design:
    [047] The animals were anesthetized by intravenous administration of ketamine and xilaxin. Three animals were used for each filling tested on day 0 = T0 - injection of the samples (1 ml of hydrogel per sample) after scraping the back of the rabbits; - measurement of swelling in all rabbits and macroscopic observation for adverse events. Day 7: T7 - Measurement of swelling volume and macroscopic observation for adverse events.
    [048] The swelling volume was calculated using the formula: (2/3 x π) x (r1) x (r2) x (r3) where: (r1), (r2) and (r3) represent the width, swelling length and height, respectively, measured with a calibration instrument. Results:
    [049] The innovative filling did not cause any inflammatory event in the treated dermis.
    [050] The results obtained for the residence time are shown in figure 1: the proportion of swelling evaluated in the first weeks of treatment (expressed as mm3) demonstrated that the gel, according to the invention is capable of inducing a greater volume of swelling in the skin than the control, remaining high even after 7 days, again to a much greater extent than the commercial filler used in comparison. This discovery clearly confirms that the new fillers immediately produce significant dermal hydration and their effect is attributed to the presence of the HYADD derivative which, due to its rheological chemical characteristics, proved to be essential for the promotion of immediate skin filling remaining stable over time. Example 11: Synthesis of HBC 500 (HA 500-730 kDa) process B
    [051] 18.75 g of sodium salt of HA with a molecular weight of 500-730 kDa, produced by fermentation are dispersed in 133 ml of a 0.25M NaOH solution containing 885 μl of BDDE. The mixture is then heated to 45 ° C for 2.5 hours. The mixture is hydrated overnight with 0.62 L of a solution containing a stoichiometric amount of HCl, 2.65 g of NaCl and 2.7 g of lidocaine hydrochloride, slowly stirring. Example 12: preparation of ACP gel: HBC 500, in the ratio of 25:75 process B
    [052] 6.25 g ACP 200 hyaluronic acid internal ester are solubilized in 250 ml of a solution containing 4.4 g of NaCl under slow stirring. When the hydration is finished, the gel is combined with the gel obtained according to example 11, in a mixer equipped with a system for mixing semi-solids, until homogeneous. The obtained gel is extruded by means of a flat membrane filter with a nominal particulate matter retention speed of 70 μm. The product thus obtained is introduced into glass syringes and sterilized in a cycle with F0 = 13 at 121.5 ° C. Example 13: preparation of HYADD: HBC 500 gel, in the ratio 25:75 process B
    [053] 6.25 g of HYADD hexadecylamide are solubilized in 250 ml of a solution containing 4.4 g of NaCl under slow stirring. When the hydration is finished, the gel is combined with the gel obtained according to example 1 in a mixer equipped with an orbital mixing system until homogeneous. The obtained gel is extruded by a flat membrane filter with a nominal particulate matter retention rate of 70 μm. The product thus obtained is introduced into glass syringes and sterilized in a cycle with F0 = 13 at 121.5 ° C. Example 14: Synthesis of HBC 500 (HA 500-730 kDa) process B
    [054] 125 g of sodium salt of HA with a molecular weight of 500 730 kDa, produced by fermentation are dispersed in 1.33 l of a 0.25 M NaOH solution containing 9.4 ml of BDDE. The mixture is heated to 45 ° C for 2.5 hours. The mix is hydrated overnight with a 6.2 l solution containing a stoichiometric amount of HCl, 26.5 g of NaCl and 27 g of lidocaine hydrochloride under slow stirring. Example 15: preparation of ACP gel: HBC 500, in the ratio 50:50 process B
    [055] 125 g of internal ester of hyaluronic acid ACP200 are solubilized in a 2.5 l solution containing 44 g of NaCl under slow stirring. When hydration is complete, the gel is combined with the gel obtained according to example 14, in a mixer equipped with an orbital system equipped with a honing and scraper. The obtained gel is extruded by a flat membrane filter with a nominal particulate material retention speed of 45 μm. The product thus obtained is introduced into glass syringes and sterilized in a cycle with F0 = 13 at 121.5 ° C. Example 16: skin filling and tolerability of ACP: HBC gel in the intradermal model of administration to rabbits
    [056] This experiment was carried out as described in Example 10, using a gel prepared as described in examples 11-12 and comparing it with the Belotero® control and with a second commercial filler, Regenyal Idea.
    [057] For this experiment, the Depositor not only determined the volume of skin swelling caused by the treatment, but also evaluated the total residence time of the gel / filler according to the invention caused by the treatment, in comparison with two well-known commercial fillers. which represent the final comparator element, since both consist of crosslinked HA with BDDE.
    [058] Skin swelling in the treated rabbits was measured biweekly (with macroscopic observation for adverse events) for a maximum of 96 days. Results:
    [059] Figure 2 shows the results obtained: the findings described above have been confirmed, that is, the immediate hydration of the treated dermis (mainly in the first 7 days) to a surprisingly greater extent than in the controls; in addition, the dimension of skin swelling became more evident and the residence time longer than that of the two commercial comparators. At the end of the experiment, the innovative filling according to the invention was still present, while the two controls had almost disappeared.
    [060] The methods described here can, of course, be modified in a number of ways. These modifications should not be considered outside the spirit and prospects of the invention; being that, all the modifications that will become evident to those of common practice in the technique are included in the scope of the attached claims.
    权利要求:
    Claims (13)
    [0001]
    1. Biomaterials, characterized by the fact that they are obtainable by mixing: - a self-crosslinked hyaluronic acid derivative (ACP) with - a cross-linked hyaluronic acid derivative (HBC) with 1,4-butanediol diglycidyl ether (BDDE), in the ratio between 10:90 and 90:10 as new fillers and / or as body shaping products.
    [0002]
    2. Biomaterials according to claim 1, characterized by the fact that the weight ratio is 90:10 to 50:50, as bio-revitalization fillers.
    [0003]
    3. Biomaterials according to claim 1, characterized by the fact that the weight ratio is from 10:90 to 50:50 with a volume-intensifying effect.
    [0004]
    4. Biomaterials according to claim 3, characterized by the fact that the weight ratio is 25:75.
    [0005]
    5. Process of mixing ACP with HBC, characterized by the fact that HBC is prepared by a process comprising the following steps: a. dissolving in alkaline solution of BDDE diepoxide in a stoichiometric ratio of 2.5 to 25 mole% of the repeat units of hyaluronic acid, followed by b. dispersion of hyaluronic acid (HA) in the solution referred to in paragraph a) at room temperature; ç. triggering of the reaction by thermal activation of the solution referred to in paragraph b) being heated to a temperature between 35 and 55 ° C between 2 and 36 hours; d. extrusion of the mass obtained through a metal sieve, in order to reduce it to particles with a size of approximately 600 μm; and. hydration of the gel obtained by diluting it with water by a factor of 3 to 20; f. pH correction to neutral with an aqueous HCl solution; g. precipitation with a water-soluble organic solvent until the product is obtained in powder form; H. washing with organic solvents containing water; i. vacuum drying until residual solvents under 400 ppm have been eliminated and an HBC powder is obtained; wherein the mixing process comprises the following steps: j. mixing ACP powder with HBC powder in the ratio of ACP: HBC between 90:10 and 10:90; k. hydration with saline or phosphate buffer, leading to a concentration of total HA between 12 and 27 mg / ml; l. extrusion at a temperature between 25 and 65 ° C through a sieve with a mesh between 50 and 500 μm; m. syringe filling; n. hot sterilization from saturated steam at a temperature between 120 and 124 ° C for at least 10 minutes.
    [0006]
    6. Process of mixing ACP with HBC, in which HBC is prepared by a process comprising the following steps: a. dissolving in alkaline solution of BDDE diepoxide in a stoichiometric ratio of 2.5 to 25 mole% of the repeat units of hyaluronic acid, followed by b. dispersion of HA in the solution referred to in paragraph a), at room temperature; ç. triggering of the reaction by thermal activation of the solution referred to in paragraph b) being heated to a temperature between 35 and 55 ° C between 2 and 36 hours; d. pH correction to neutral with an aqueous HCl solution; and. hydration of the gel obtained by diluting it with water by a factor of 3 to 20; wherein the mixing process comprises the following steps: f. mixing ACP in gel or powder with HBC in gel form in an ACP: HBC ratio between 90:10 and 10:90; g. crushing and homogenization passing through a filter with a retention coefficient of particulate matter between 25 and 150 μm; H. syringe filling; i. heat sterilization with saturated steam at a temperature between 120 and 124 ° C for at least 10 minutes.
    [0007]
    Process according to claim 5 or 6, characterized in that the HA used for the preparation of the HBC and ACP derivatives has an average molecular weight between 400 and 3 x 106 Da, preferably between 1 x 105 Da and 1 x 106 Da, and more preferably between 200,000 and 1 x 106 Da.
    [0008]
    8. Process according to claim 5, characterized by the fact that the water-soluble organic solvent is selected from ethanol, methanol, isopropanol, n-propanol, dioxane, acetonitrile, acetone and / or mixtures thereof.
    [0009]
    9. Biomaterials, characterized by the fact that they are obtained by the process as defined in claim 6, in which the weight ratio of ACP: HBC is 25:75.
    [0010]
    10. Biomaterials according to any of the preceding claims, characterized by the fact that the vehicle is saline.
    [0011]
    11. Biomaterials according to any of the preceding claims, characterized by the fact that they contain lidocaine.
    [0012]
    12. Biomaterials according to claim 9, characterized by the fact that they contain lidocaine, in which the vehicle consists of saline solution.
    [0013]
    13. Use of biomaterials as defined in any of the preceding claims, characterized by the fact that they are as new fillers and / or as new products for body shaping, in the treatment of skin imperfections, in dermatology, in dermocosmetology and / or in surgery aesthetics.
    类似技术:
    公开号 | 公开日 | 专利标题
    BR112012004157B1|2021-03-09|biomaterials, process of mixing acp with hbc and using them
    ES2556475T3|2016-01-18|Injectable composition based on hyaluronic acid, polyol | and lidocaine, heat sterilized
    TWI641393B|2018-11-21|Sterile injectable aqueous formulation containing cross-linked hyaluronic acid and hydroxyapatite for cosmetic use
    BR112015031026B1|2020-07-07|method for crosslinking hyaluronic acid or one of its salts or other biocompatible polymers; method for preparing an injectable hydrogel; hydrogel; kit and use
    BRPI9609534B8|2020-05-12|process for preparing a cross-linked biocompatible polysaccharide gel composition, and polysaccharide gel composition.
    AU2013276844B2|2017-09-28|Method of preparing a composition based on hyaluronic acid
    US7521434B2|2009-04-21|Cross-linked gels of hyaluronic acid with hydrophobic polymers and processes for making them
    BR112013012772B1|2019-05-14|Preparation and / or formulation of polysaccharide crosslinked proteins
    ES2637202T3|2017-10-11|Hyaluronic acid based formulations
    RU2477138C1|2013-03-10|Method for preparing filling material for plastic surgery and instrumental cosmetology, filling material and method for introducing filling material into problem area
    BR112020024637A2|2021-03-02|stabilized hyaluronic acid
    WO2017001057A1|2017-01-05|Method of preparing a composition based on hyaluronic acid
    KR102132478B1|2020-07-10|Hyaluronic Acid-Polydeoxyribonucleotide Complex and Its Film and Preparation Method Thereof
    TW202042848A|2020-12-01|Cosmetic method for filling tissue and use of chitosan solution for manufacturing a composition used in a method of treatment by surgery
    BR112021011574A2|2021-08-31|HYALURONIC ACID MODIFIED BY ALDEHYDE, METHOD FOR PREPARING IT AND APPLICATIONS THEREOF
    同族专利:
    公开号 | 公开日
    EP2470230A2|2012-07-04|
    RU2692800C2|2019-06-27|
    ITPD20090246A1|2011-02-28|
    AU2010288937B2|2015-03-05|
    WO2011023355A3|2011-10-27|
    ES2509225T3|2014-10-17|
    EP2772273B1|2015-12-02|
    CA2995957A1|2011-03-03|
    CY1115628T1|2017-01-04|
    DK2470230T3|2014-10-06|
    BR112012004157B8|2021-06-22|
    CA2772142A1|2011-03-03|
    EP2772273B9|2016-05-18|
    RU2543329C2|2015-02-27|
    RU2012106800A|2013-08-27|
    KR101749660B1|2017-06-21|
    PL2470230T3|2015-02-27|
    SI2470230T1|2014-11-28|
    HRP20140981T2|2016-05-06|
    CY1117207T1|2017-04-05|
    CA2995957C|2020-04-28|
    US8846640B2|2014-09-30|
    ES2563160T3|2016-03-11|
    CA2772142C|2018-04-10|
    HK1172850A1|2013-05-03|
    HK1196787A1|2014-12-24|
    SMT201600062B|2016-04-29|
    JP2013502941A|2013-01-31|
    BR112012004157A2|2020-08-11|
    RU2014149799A3|2018-08-28|
    SI2772273T1|2016-04-29|
    HRP20140981T8|2015-01-16|
    CN102548590A|2012-07-04|
    JP5976537B2|2016-08-23|
    DK2772273T3|2016-03-07|
    AU2010288937A1|2012-03-15|
    HUE026830T2|2016-07-28|
    CN104623730B|2016-11-16|
    US20120190644A1|2012-07-26|
    EP2772273A3|2014-12-03|
    KR20120107455A|2012-10-02|
    HRP20140981T1|2014-11-21|
    HRP20160196T1|2016-03-25|
    EP2772273A2|2014-09-03|
    PL2772273T3|2016-05-31|
    IT1395392B1|2012-09-14|
    MX2012002364A|2012-03-29|
    RU2014149799A|2016-07-10|
    EP2470230B1|2014-07-23|
    PT2470230E|2014-10-07|
    CN102548590B|2015-02-25|
    CN104623730A|2015-05-20|
    WO2011023355A2|2011-03-03|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    SE8501022L|1985-03-01|1986-09-02|Pharmacia Ab|FORMAT CREATES AND PROCEDURES FOR ITS PREPARATION|
    IT1198449B|1986-10-13|1988-12-21|F I D I Farmaceutici Italiani|ESTERS OF POLYVALENT ALCOHOLS OF HYALURONIC ACID|
    IT1219587B|1988-05-13|1990-05-18|Fidia Farmaceutici|SELF-CROSS-LINKED CARBOXYLY POLYSACCHARIDES|
    US5827937A|1995-07-17|1998-10-27|Q Med Ab|Polysaccharide gel composition|
    IT1288290B1|1996-06-21|1998-09-11|Fidia Spa In Amministrazione S|SELF-LETICULATED HYALURONIC ACID AND RELATED PHARMACEUTICAL COMPOSITIONS FOR THE TREATMENT OF ARTHROPATHIES|
    IT1296689B1|1997-11-06|1999-07-14|Fidia Advanced Biopolymers Srl|FOREIGN DERIVATIVES OF HYALURONIC ACID WITH VISCOELASTIC PROPERTIES AND THEIR USE IN THE BIOMEDICAL-SANITARY FIELD|
    ITPD980169A1|1998-07-06|2000-01-06|Fidia Advanced Biopolymers Srl|AMIDES OF HYALURONIC ACID AND ITS DERIVATIVES AND PROCESS FOR THEIR PREPARATION.|
    IT1303735B1|1998-11-11|2001-02-23|Falorni Italia Farmaceutici S|CROSS-LINKED HYALURONIC ACIDS AND THEIR MEDICAL USES.|
    IT1317358B1|2000-08-31|2003-06-16|Fidia Advanced Biopolymers Srl|CROSS-LINKATED DERIVATIVES OF HYALURONIC ACID.|
    WO2005112888A2|2004-05-20|2005-12-01|Mentor Corporation|Methods for making injectable polymer hydrogels|
    ITPD20050056A1|2005-03-02|2006-09-03|Fidia Farmaceutici|AMIDID DERIVATIVES OF HYALURONIC ACID IN OSTEOARTROSI|
    ITPD20050146A1|2005-05-20|2006-11-21|Fidia Farmaceutici|RE-ABSORBABLE FILLERS CONSISTING OF LIPOSOMAS AND HYALURONIC ACID AND OR ITS DERIVATIVES|
    FR2909560B1|2006-12-06|2012-12-28|Fabre Pierre Dermo Cosmetique|HYALURONIC ACID GEL FOR INTRADERMAL INJECTION|
    JP5165281B2|2007-06-01|2013-03-21|株式会社バイオベルデ|Two-reactor type water-containing medical gel-forming agent and hyaluronic acid gel obtained therefrom|
    FR2918377B1|2007-07-05|2010-10-08|Estelle Piron|CO-RETICLE GEL OF POLYSACCHARIDES|
    ES2381417T3|2007-09-28|2012-05-28|Shiseido Company, Ltd.|Inflatable crosslinked hyaluronic acid powder and process for manufacturing it|
    WO2009043111A1|2007-10-04|2009-04-09|Ultraceuticals R & D Pty Ltd|Composition and method for dermal regeneration|
    US8394782B2|2007-11-30|2013-03-12|Allergan, Inc.|Polysaccharide gel formulation having increased longevity|
    FR2924615B1|2007-12-07|2010-01-22|Vivacy Lab|HYDROGEL COHESIVE BIODEGRADABLE.|
    WO2009077399A1|2007-12-19|2009-06-25|Evonik Goldschmidt Gmbh|Crosslinked hyaluronic acid in emulsion|
    US8357795B2|2008-08-04|2013-01-22|Allergan, Inc.|Hyaluronic acid-based gels including lidocaine|US8124036B1|2005-10-27|2012-02-28|ADA-ES, Inc.|Additives for mercury oxidation in coal-fired power plants|
    TR201901431T4|2007-11-16|2019-02-21|Aclaris Therapeutics Inc|Compositions and methods for treating purpura.|
    US8394782B2|2007-11-30|2013-03-12|Allergan, Inc.|Polysaccharide gel formulation having increased longevity|
    US8357795B2|2008-08-04|2013-01-22|Allergan, Inc.|Hyaluronic acid-based gels including lidocaine|
    EP2324064B1|2008-09-02|2017-11-08|Tautona Group LP|Threads of hyaluronic acid and/or derivatives thereof, methods of making thereof and uses thereof|
    US20110172180A1|2010-01-13|2011-07-14|Allergan Industrie. Sas|Heat stable hyaluronic acid compositions for dermatological use|
    US8496894B2|2010-02-04|2013-07-30|ADA-ES, Inc.|Method and system for controlling mercury emissions from coal-fired thermal processes|
    CA2788820C|2010-02-04|2021-09-21|Michael Durham|Method and system for controlling mercury emissions from coal-fired thermal processes|
    US8784757B2|2010-03-10|2014-07-22|ADA-ES, Inc.|Air treatment process for dilute phase injection of dry alkaline materials|
    US8383071B2|2010-03-10|2013-02-26|Ada Environmental Solutions, Llc|Process for dilute phase injection of dry alkaline materials|
    FR2957254B1|2010-03-12|2016-10-14|Allergan Ind Sas|FLUID COMPOSITIONS FOR IMPROVING SKIN CONDITIONS|
    EP2550027B2|2010-03-22|2019-03-20|Allergan, Inc.|Polysaccharide and protein-polysaccharide cross-linked hydrogels for soft tissue augmentation|
    US9005605B2|2010-08-19|2015-04-14|Allergan, Inc.|Compositions and soft tissue replacement methods|
    US8889123B2|2010-08-19|2014-11-18|Allergan, Inc.|Compositions and soft tissue replacement methods|
    US8883139B2|2010-08-19|2014-11-11|Allergan Inc.|Compositions and soft tissue replacement methods|
    KR102015676B1|2011-06-03|2019-10-21|알러간, 인코포레이티드|Dermal filler compositions including antioxidants|
    US9408797B2|2011-06-03|2016-08-09|Allergan, Inc.|Dermal filler compositions for fine line treatment|
    US20130096081A1|2011-06-03|2013-04-18|Allergan, Inc.|Dermal filler compositions|
    US9393263B2|2011-06-03|2016-07-19|Allergan, Inc.|Dermal filler compositions including antioxidants|
    HRP20211659T1|2011-08-10|2022-02-04|Glycores 2000 Srl|Degradation-resistant cross-linked, low-molecular-weight hyaluronate|
    CN104105474B|2011-09-14|2018-04-06|阿勒根公司|Dermal augmentation agent composition for microgroove treatment|
    US20130244943A1|2011-09-06|2013-09-19|Allergan, Inc.|Hyaluronic acid-collagen matrices for dermal filling and volumizing applications|
    KR101428145B1|2011-11-24|2014-08-08|아모레퍼시픽|Water-insoluble gel composition and manufacturing method of the same|
    ITPD20120098A1|2012-03-30|2013-10-01|Fidia Farmaceutici|"NEW PHARAMACEUTICAL FORMULATIONS CONTAINING CONDROITIN SULFATE AND DERIVATIVES OF HYALURONIC ACID"|
    ITMI20120732A1|2012-05-03|2013-11-04|B S Srl|POLYSACCHARIDES RETICULATED IN SHAPE MEMORY|
    RU2640911C2|2012-06-15|2018-01-12|Мерц Фарма Гмбх Унд Ко. Кгаа|Method for obtaining of composition based on hyaluronic acid|
    FR2997085B1|2012-10-24|2015-05-29|Teoxane|PROCESS FOR PREPARING RETICULATED GEL|
    ITPD20120360A1|2012-11-30|2014-05-31|Fidia Farmaceutici|"NEW VISCOELASTIC GELES IN OPHTHALMIC SURGERY"|
    US20140315828A1|2013-04-22|2014-10-23|Allergan, Inc.|Cross-linked silk-hyaluronic acid compositions|
    ITPD20130110A1|2013-04-24|2014-10-25|Fidia Farmaceutici|PHARMACEUTICAL COMPOSITIONS INCLUDING HYALURONIC ACID FOR THE TREATMENT OF BLACK DISC|
    CA2916330A1|2013-06-28|2014-12-31|Galderma S.A.|A process for preparing a cross-linked hyaluronic acid product|
    CA2917715A1|2013-07-08|2015-01-15|Denka Company Limited|Core-shell crosslinked hyaluronic acid gel particles, production method for same, and medical material|
    EP3035903B1|2013-08-20|2018-08-22|Anutra Medical, Inc.|Syringe fill system and method|
    USD774182S1|2014-06-06|2016-12-13|Anutra Medical, Inc.|Anesthetic delivery device|
    USD763433S1|2014-06-06|2016-08-09|Anutra Medical, Inc.|Delivery system cassette|
    USD750768S1|2014-06-06|2016-03-01|Anutra Medical, Inc.|Fluid administration syringe|
    EP3620184A1|2014-09-30|2020-03-11|Allergan Industrie, SAS|Stable hydrogel compositions including additives|
    FR3029928B1|2014-12-15|2018-03-09|Teoxane|RETICULATED GEL|
    RU2610006C1|2016-03-23|2017-02-07|Наталья Павловна Михайлова|Lypotherapy method|
    AU2017315431B2|2016-08-24|2020-06-25|Allergan, Inc.|Co-crosslinked hyaluronic acid-silk fibroin hydrogels for improving tissue graft viability and for soft tissue augmentation|
    US20210283172A1|2017-04-12|2021-09-16|Urigen Pharmaceuticals, Inc.|Article of manufacture comprising local anesthetic, buffer, and glycosaminoglycan in syringe with improved stability|
    RU2020123793A|2017-12-22|2022-01-25|Нестле Скин Хелс Са|INJECTION GEL PRODUCT|
    IT201800001890A1|2018-01-25|2019-07-25|Fidia Farm Spa|PHARMACEUTICAL COMPOSITIONS FOR THE TREATMENT OF POSTOPERATIVE PAIN|
    WO2020184744A1|2019-03-08|2020-09-17|주식회사 지씨에스|Method for producing complex|
    RU2746877C1|2020-07-23|2021-04-21|Роман Дмитриевич Лебедев|Porous filter element and its manufacturing method|
    法律状态:
    2020-08-18| B06U| Preliminary requirement: requests with searches performed by other patent offices: procedure suspended [chapter 6.21 patent gazette]|
    2021-01-12| B09A| Decision: intention to grant [chapter 9.1 patent gazette]|
    2021-03-09| B16A| Patent or certificate of addition of invention granted|Free format text: PRAZO DE VALIDADE: 10 (DEZ) ANOS CONTADOS A PARTIR DE 09/03/2021, OBSERVADAS AS CONDICOES LEGAIS. |
    2021-06-22| B16C| Correction of notification of the grant|Free format text: PRAZO DE VALIDADE: 20 (VINTE) ANOS CONTADOS A PARTIR DE 25/08/2010, OBSERVADAS AS CONDICOES LEGAIS. PATENTE CONCEDIDA CONFORME ADI 5.529/DF, QUE DETERMINA A ALTERACAO DO PRAZO DE CONCESSAO |
    优先权:
    申请号 | 申请日 | 专利标题
    ITPD2009A000246A|IT1395392B1|2009-08-27|2009-08-27|VISCOELASTIC FROSTS LIKE NEW FILLERS|
    ITPD2009A000246|2009-08-27|
    PCT/EP2010/005161|WO2011023355A2|2009-08-27|2010-08-25|Viscoelastic gels as novel fillers|
    [返回顶部]