WO2019009823A1 - A pharmaceutical composition on containing atorvastatin - Google Patents

A pharmaceutical composition on containing atorvastatin Download PDF

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Publication number
WO2019009823A1
WO2019009823A1 PCT/TR2017/050299 TR2017050299W WO2019009823A1 WO 2019009823 A1 WO2019009823 A1 WO 2019009823A1 TR 2017050299 W TR2017050299 W TR 2017050299W WO 2019009823 A1 WO2019009823 A1 WO 2019009823A1
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Prior art keywords
polymer
atorvastatin
active agent
chitosan
mixture
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PCT/TR2017/050299
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French (fr)
Inventor
Sevda SENEL
Ayben Isilay OZDOGAN
Gulcin AKCA
Kemal KOSEMEHMETOGLU
Yagmur Deniz ILARSLAN
Alper Bektas ISKIT
Original Assignee
Senel Sevda
Ozdogan Ayben Isilay
Akca Gulcin
Kosemehmetoglu Kemal
Ilarslan Yagmur Deniz
Iskit Alper Bektas
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Application filed by Senel Sevda, Ozdogan Ayben Isilay, Akca Gulcin, Kosemehmetoglu Kemal, Ilarslan Yagmur Deniz, Iskit Alper Bektas filed Critical Senel Sevda
Priority to PCT/TR2017/050299 priority Critical patent/WO2019009823A1/en
Publication of WO2019009823A1 publication Critical patent/WO2019009823A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0053—Mouth and digestive tract, i.e. intraoral and peroral administration
    • A61K9/0063—Periodont
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
    • A61K9/146—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with organic macromolecular compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/02—Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis

Definitions

  • the present invention relates to a pharmaceutical composition containing atorvastatin, which is an active agent belonging to the statin group which has an anti-inflammatory effect, and/or a pharmaceutically acceptable salt thereof and at least one carrier polymer; and is developed in order to be used in the treatment of periodontal diseases. Thanks to this locally administered composition; the active agent with improved solubility is combined with the chitosan gels used as the carrier system to obtain high anti-inflammatory effect.
  • Periodontal diseases are inflammatory diseases which affect the gingiva and the other tissues supporting the teeth. Periodontal diseases start with gingivitis. Periodontitis is a more advanced stage of periodontal diseases. Alveolar bone is damaged in addition to the other tissues supporting the teeth. "Periodontal pockets" occur between the teeth and the gingiva. The presence of periodontal pockets facilitates the settling of the infection and the progression of the diseases. As the diseases progresses, the teeth start to become loose and this may lead to extraction. Thinning of the alveolar bone, degeneration of the collagen fibers, loss of periodontal ligaments and loss of attachment are the other outcomes of the disease.
  • the first treatment in periodontal diseases is the scaling of bacterial plaque and tartar which cause the disease to progress.
  • mechanical practices such as tooth surface cleaning and root planning are performed.
  • these practices are limited to only being the first line treatment in periodontal diseases and the bacteria which cannot be removed or accessed due to their attachment to the periodontal tissues cause the disease to progress and lead to loss of attachment and deterioration of the alveolar bone and the periodontal ligament.
  • antibiotics such as tetracycline, metronidazole, clindamycin, amoxicillin, penicillin and cephalosporin are administered in systemic manner for treatment. Feres et al. mention the systemic use of antibiotics in their article for the treatment of periodontal disease ⁇ Systemic antibiotics in the treatment of periodontitis, Periodontology 2000, 67(1), 131-186, 2015). However, as the use of antibiotics cause severe bacterial resistance, the use of antibiotics to which the pathogens are resistant fails in stopping the progression of the disease.
  • systemic drugs for treatment causes the drug to suffer the hepatic first pass effect, to undergo enzymatic or hydrolytic decomposition in the gastrointestinal tract and consequently to have a lower amount of drug that reaches the area of effect.
  • drugs with higher dose are used in this practice, which is undesirable for human health.
  • Topical chemical practices also exist in addition to the mechanical cleaning performed to control and prevent the formation of plaque.
  • Mouthwash solutions are frequently used in the topical practice.
  • chlorhexidine which an antimicrobial agent is proven to inhibit dental plaque.
  • these solutions have certain disadvantages such as the necessity to use them several times a day, and the inability of the solution to stay at the area of effect for long period of time due to the lack of adhesion to the periodontal pocket occurring in periodontal disease and disposal by saliva.
  • Local drug treatment outclasses systemic drug treatment thanks to advantages such as high drug concentration at the area of administration, low side effects, non-necessity of daily administration.
  • bioadhesive drug formulations are required for preventing the removal of the locally administered drug from the area of administration due to the presence of saliva and tongue movement, and for ensuring that said drug stays at the area long enough to create the desired effect.
  • Statins are drugs which are used to treat hypercholesterolemia in atherosclerosis and cardiovascular system disorders, and they regulate the cholesterol level in blood by inhibiting the biosynthesis of cholesterol in the liver. They reduce the levels of low density lipoprotein (LDL) in the plasma by acting as 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG CoA reductase) inhibitors.
  • LDL low density lipoprotein
  • HMG CoA reductase 3-hydroxy-3-methyl-glutaryl-CoA reductase
  • various drugs belonging to the statin group such as atorvastatin, simvastatin, rosuvastatin, lovastatin, fluvastatin and pravastatin.
  • statins exhibit pleiotropic effects. "Pleiotropic effect” is the effect exhibited by the drug apart from the effect it is specifically developed for. Besides their lipid lowering effects, statins have anabolic effect and anti-inflammatory effect on bones as their pleiotropic effects. In the known art, there are certain studies showing that the use of statin reduces the risk of inflammation [1, 2].
  • Patent document no: US 2001006656 Al describes an aerosol formulation which contains HMG- CoA reductase inhibitor and which is used for spray administration. Said HMG-CoA reductase inhibitor can be lovastatin, pravastatin, simvastatin, cerivastatin, fluvastatin, atorvastatin or mevastatin.
  • the invention is developed in order to be used for the treatment of respiratory system diseases related to inflammation such as asthma, pneumonia and chronic bronchitis.
  • IL-6 interleukin-6
  • IL-8 interleukin-8
  • IL-10 interleukin-10
  • IL- lbeta interleukin-lbeta
  • Atorvastatin is a drug with anti-inflammatory effects which belongs to the statin group and has low water solubility. It does not dissolve in aqueous solutions at and below pH 4; and it slightly dissolves in distilled water and pH 7.4 phosphate buffer.
  • the chemical formula of Atorvastatin, the chemical name of which is [(3R,5R)-7-[2-(4-fluorophenyl)-3-phenyl-4-(phenylcarbamoyl)-5- propane-2-ylpyrol-l-yl]-3,5-dihydroxyheptanoic acid] is shown below:
  • Physical modifications are processes such as decreasing particle sizes via micronisation, changing crystal properties, creating polymorphs, creating complex with cyclodextrin, solutionizing, preparing the eutectic mixture of the drug inside the carrier and preparing the solid dispersion thereof.
  • Chemical modifications are processes such as creating a soluble prodrug and creating the salts of the drug.
  • Chitosan is a natural polymer obtained by the partial deacetylation chitin which is present in the shells of shellfish such as crabs and shrimps, shells of insects and cell walls of fungi.
  • chitosan which is a cationic and biodegradable polymer, change depending on properties such as the level of deacetylation and molecular weight.
  • chitosan being a biocompatible, biodegradable, non-toxic and non-allergenic natural polymer; it has antimicrobial, hemostatic, wound healing and tissue regenerating biological effects.
  • chitosan since it is a bioadhesive polymer, it is typically preferred in carrier systems which are locally administered [4].
  • the structure of chitosan which is a copolymer of D- glucosamine and N-acetyl-D-glucosamine, is presented below.
  • atorvastatin as an antiinflammatory drug in the periodontal disease is known.
  • solubility of atorvastatin of the invention is increased and atorvastatin is used inside a chitosan based gel in order to be administered locally in the treatment of the periodontal disease.
  • the solubility problem of atorvastatin in locally administered pharmaceutical compositions developed for the treatment of the periodontal disease is eliminated. Furthermore, higher effects can be achieved with lower doses thanks to the local administration of the invention, and the drug is not easily removed by the presence of saliva and/or tongue movements, thereby enabling the drug to have an effect on the area of administration for a long period of time. Thus, bioavailability increases, the frequency of side effects decreases and patient compatibility rises. Also, there are various advantages such as ease of administration and prolonged presence at the area of administration due to the bioadhesive properties of the drug.
  • the present invention relates to a new pharmaceutical composition developed by eliminating the solubility problem of atorvastatin, which belongs to the statin group, through various pharmaceutical processes in order to be used in the treatment of periodontal diseases.
  • local drug treatment outclasses systemic drug treatment thanks to advantages such as high drug concentration, low side effects, non-necessity of daily administration.
  • Figure 1 1 . IL-6 levels 4 hours after administration in hGF cell culture in (1/2) dilution of formulations
  • FIG. Histological micrograph of the periodontal tissues of the healthy control group ((-) control) (at the end of Week 1)
  • FIG. 20 Histological micrograph of the periodontal tissues of the control group with periodontal disease without administration ((+) control) (at the end of Week 1)
  • FIG 21 Histological micrograph of the periodontal tissues of the group administered with Atorvastatin (AT) (at the end of Week 1)
  • FIG 22 Histological micrograph of the periodontal tissues of the group administered with base chitosan gel containing Atorvastatin (gK-AT) (at the end of Week 1)
  • FIG 23 Histological micrograph of the periodontal tissues of the group administered with water soluble chitosan gel containing Atorvastatin (gP-AT) (at the end of Week 1)
  • FIG. 24 Atorvastatin prepared by the melting method: Histological micrograph of the periodontal tissues of the group administered with base chitosan gel containing PEG 6000 (1: 12) solid dispersion (gK ⁇ ML-dPEG12 - AT) (at the end of Week 1) Detailed Description of the I nvention
  • the present invention relates to a gel, fiber, sponge, film, micro or nanoparticle system administered locally with an active agent selected from the statin group and/or a pharmaceutically acceptable salt thereof and at least one carrier polymer for use in the treatment of the periodontal disease, and the production method of said pharmaceutical composition.
  • the active agent is selected among atorvastatin, simvastatin, rosuvastatin, lovastatin, fluvastatin, pravastatin from the statin group and/or the acceptable salts thereof.
  • the preferred form of the invention is a gel composition of atorvastatin and/or and acceptable salt thereof. Atorvastatin which is used in the gel composition is selected within the range of 0.5-4% (weight/volume, w/v).
  • Atorvastatin and/or an acceptable salt thereof are used preferably as 2% (w/v) in the gel composition.
  • Atorvastatin which is known to exhibit properties regulating the cholesterol level and reducing lipids when used in high doses, exhibits an anti-inflammatory effect when used in low doses in our study.
  • the drugs used in high doses damage the kidneys and cause renal toxicity, it is important to choose a treatment dosage where the drug exhibits an effect in low doses.
  • the carrier polymer used in the preparation of the gel composition is selected among polymers with bioadhesive properties, such as chitosans which have various molecular weights (MW), various deacetylation levels and various solubility, sodium alginate, hyraluronic acid, pectin, agarose, gelatin, carrageenan, aminodextran, hydroxypropyl cellulose (HPC), polyethylene glycol (PEG), polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP), ethyl cellulose, hydroxy ethyl cellulose, polycarbophil, methylvinyl ether and methacrylic acid copolymer, polyacrylates, polyalkylcyanoacrylate, acrylic acid and ehtlyhexylacyrlate copolymer, polymethacrylate, carbopol, methyl hydroxy ethyl cellulose, methyl cellulose, hydroxy propyl methyl cellulose (HPMC) and carboxy methyl
  • atorvastatin with increased solubility is used and as the carrier polymer, base chitosan (dissolves in diluted acidic solutions) which is a biodegradable, biocompatible, natural, non-toxic polymer or water soluble chitosan containing different salts thereof, is used.
  • Chitosans are preferred in the formulation due to the antimicrobial, anti-inflammatory, tissue regenerating, wound and bone healing etc. bioactive properties they possess. Also, the physical-chemical and bioactive properties of chitosans with various molecular weights (MW), various deacetylation levels and various solubility vary.
  • the chitosan used in the formulation has a high anti-inflammatory effect.
  • the anti-inflammatory activity of the drug increases thanks to the base chitosan and water soluble chitosan used in the present invention.
  • chitosan with 85% deacetylation level is used as base chitosan and chloride salt of chitosan with 86% deacetylation level is used as water soluble chitosan.
  • Chitosan can be used in the range of 0.1% - 10% (w/v) in the formulation.
  • the chitosan used in the invention is at the rate of 2% (w/v).
  • Base chitosan dissolves in diluted acidic solutions.
  • organic acids such as acetic acid, lactic acid, citric acid, malic acid and formic acid are used in the invention.
  • These acid solutions can be prepared as 0.1% - 5%.
  • 1% diluted lactic acid solution is prepared by using diluted lactic acid.
  • the gel formulation to be prepared with 2% chitosan which exhibits properties of pseudoplastic flow and high bioadhesion can be obtained by preparing the lactic acid solution as 1%.
  • release studies have been performed with only atorvastatin, and the mixture thereof with base chitosan and water soluble chitosan gels. Release studies are performed at 37°C by using Franz diffusion cells with a 2.01 cm 2 diffusion area and 20 mL receptor volume. pH 6.8 phosphate buffer is used as the diffusion medium. 1 mL samples are collected at certain time intervals and the same amount of buffer is added instead. The amount of active agent in the collected samples is measured spectrophotometrically. Figure 1 shows the graph for the release studies. Atorvastatin is dissolved in PEG400. Release is found to be low when the active agent concentrations released from the gels are compared to this PEG400-AT mixture.
  • Solubility of atorvastatin needs to be increased in order to improve the release profile.
  • the method of creating solid dispersion is utilized in order to increase the solubility of atorvastatin.
  • the method of creating solid dispersion is a more practical approach for increasing solubility as it is easier and more feasible than chemical modifications. Thanks to the method of creating solid dispersion, particle size is reduced, wettability and porosity of the particles increase, drugs with crystal structures which have low water solubility are transformed into amorphous form and thus, solubility increases. Also, the bioavailability of the drugs increases thanks to the method of creating solid dispersion.
  • Methods of melting, hot melt extrusion, co-evaporation, rotating evaporator, spray drying, freeze drying, spray freeze drying, supercritical antisolvent, co-precipitation, electrostatic spinning, co-milling and kneading are used in order to create solid dispersions.
  • melting, co-evaporation, co-milling and kneading are used as the methods for creating solid dispersions.
  • degradation of the drug and the polymer may occur.
  • loss of drug may occur due to the drug and polymer sticking to the walls of the tools used in creating solid dispersions. Due to these disadvantages; methods of co-evaporation, melting, co-milling and kneading, which do not require heat or which operate at low temperatures, are preferred and no drug loss occurs. Additionally, these methods are preferred in the invention as it is easier to prepare solid dispersions with these methods compared to others.
  • the polymers used in creating solid dispersions can be selected among: cellulose derivatives such as hydroxypropyl methyl cellulose (HPMC), ethyl cellulose, hydroxypropyl cellulose (HPC), sodium carboxymethyl cellulose (NaCMC), starch derivatives, polyvinyl pyrrolidone, eudragit, polyethylene glycols, polymethacrylates, chitosans with various deacetylation, various molecular weights and various solubility, inulin, compritol 888 ATO, gelucire, pluronics, gelatin, cyclodextrins, Kollidon, Avicel 200, Synperonic and Plasdone.
  • HPMC hydroxypropyl methyl cellulose
  • HPC hydroxypropyl cellulose
  • NaCMC sodium carboxymethyl cellulose
  • starch derivatives polyvinyl pyrrolidone, eudragit, polyethylene glycols, polymethacrylates, chitosans with various de
  • the water solubility of the drug with crystal structure is increased via the method of creating solid dispersions.
  • Polyethylene glycol, pluronic and chitosan polymers are used in the invention for creating solid dispersions.
  • polyethylene glycol 6000 (PEG 6000), pluronic F-68 (PL F- 68) and base chitosan polymers are used.
  • the reason for using polyethylene glycol 6000 and pluronic F-68 among the water soluble polymers is that it is more effective to utilize polymers with high water solubility in order to increase the water solubility of the drug.
  • the reason for using chitosan is to ensure that the anti-inflammatory properties of the drug are further increased by making use of the bioactive properties of chitosan. As solid dispersions cannot be created with water soluble chitosan, it is not included among the polymers selected for creating solid dispersions.
  • the polymers used in preparing solid dispersions can be used at the ratios of 1:0.1 - 1:25 (active agent: polymer).
  • active agent polymer
  • the obtained values of solubility, release and dissolution change depending on the various polymers, various active agent: polymer ratios, various methods and various active agents used in the solid dispersions. For this reason, it is not possible to generalize by saying that a solid dispersion with higher solubility, release and dissolution values is going to be obtained due to increased or decreased polymer ratios.
  • solid dispersions of piroxicam with PVP K-30 have been prepared at the active agent: polymer ratios of 1:0.5, 1: 1, 1:2, 1:3, 1:4, 1:5 and 1:6; and the highest dissolution value has been obtained with the solid dispersion prepared at the ratio of 1:4 [5].
  • solid dispersions of atorvastatin with the ratios of 1: 1, 1:3, 1:9 and 1: 12 active agent: polymer
  • These prepared solid dispersions have been mixed with chitosan gels and the release studies of these compositions have been performed.
  • gKN-dK9-AT AT Base chitosan (1:9) (kneading) + 1% lactic acid solution
  • gK-CE-dPL12-AT AT:Pluronic F-68 (1: 12) (co-evaporation) + base chitosan gel gKN-dK12-AT AT:Base chitosan (1: 12) (kneading) + 1% lactic acid solution gCG-dK12-AT AT: Base chitosan (1: 12) (co-milling) + 1% lactic acid solution gP-CE-dPL12-AT AT: PEG 6000 (1: 12) (co-evaporation) + water soluble
  • Figures 2-4 show the active agent release graphs of chitosan gels containing atorvastatin solid dispersions. Release graphs are presented separately according to the type of the used polymer. It can be seen that the active agent release from the chitosan gel compositions containing solid dispersions prepared at ratios of 1: 1 and 1:3 (active agent: polymer) are low.
  • the active agent release from the chitosan gel compositions containing solid dispersions prepared at ratios of 1:9 and 1: 12 are higher than the release of atorvastatin from base and water soluble chitosan gels (gK-AT and gP-AT) in Figure 1.
  • ratios of 1:9 and 1: 12 are preferred over the ratios of 1: 1 and 1:3 (active agent: polymer) in the invention.
  • the table below shows the solid dispersions prepared at the active agent:polymer ratios of 1:9 and 1: 12 with polymers and various preparation methods.
  • Drug release from the gels of the solid dispersions prepared with base chitosan decreases when the methods of melting and co-evaporation are used, and said release from gels increases when the methods of co-milling and kneading are used. No increase in the drug solubility is observed when the methods of co-milling and kneading are used with PEG 6000.
  • solid dispersions cannot be created due to the properties of the polymer when the methods of co- milling and kneading are used with Pluronic F-68.
  • atorvastatin and the polymer are dissolved in an organic solvent, preferably methanol, and afterwards the solvent is evaporated.
  • Methanol is chosen as the solvent because atorvastatin easily dissolved in methanol.
  • methanol causes the experiment to yield incorrect results by causing cells to die in the cell culture studies carried out to show the anti-inflammatory effects of the composition; there should be no traces of methanol in the prepared composition. Therefore, methanol is preferably evaporated in the rotating evaporator at 30°C-80°C.
  • PEG 6000- atorvastatin and Pluronic F-68-atorvastatin mixtures are dried until no moisture remains. Preferably, they are kept inside the desiccator or the refrigerator for 24 hours. Afterwards, both mixtures are pulverized separately.
  • atorvastatinxhitosan solid dispersion with the co-milling method physical atorvastatin and polymer mixtures with the ratios of 1:9 and 1: 12 are mixed inside the ball mill and pulverized.
  • the mixing process is carried out for 1-24 hours at 100-600 rpm.
  • the powder mixture is preferably prepared by mixing for 1 hour at 300 rpm. The reason for choosing these conditions is that mixture sticks to the inner walls or the balls of the device when mixed at a higher speed or for a longer time. Therefore, a speed is selected where the highest yield is achieved in the shortest time.
  • atorvastatin chitosan solid dispersion with the kneading method; physical atorvastatin and polymer mixtures with the ratios of 1:9 and 1: 12 are placed inside the porcelain mortar and are mixed with organic solvent or organic solvent: water mixture, the amount of which is 1 to 10 times amount the weighed powder.
  • the organic solvent: water mixture to be used is selected from the range of 0.1: 1 - 20: 1.
  • the obtained mixture is dried inside a dryer at a temperature selected between 10°C - 80°C by keeping said mixture therein for 1 - 5 days.
  • the mixture of atorvastatin: polymer is mixed by wetting it with a methanokwater (6: 1) mixture 1.5 times the amount of the weighed powder.
  • the water solubility of atorvastatin is quite low and it can easily dissolve in methanol, it is important to obtain a solution mixture prepared at the correct ratios. Furthermore, since methanol may cause the experiment to not yield the correct results by causing cells to die in the cell culture studies carried out to show the anti-inflammatory effects of the composition; there should be no traces of methanol in the prepared composition. In other words, the methanol to be used should be present in an amount that will allow atorvastatin to dissolve and it should be used minimally in a way that it does not remain in the formulation after drying. For this reason, the obtained mixture is kept inside the oven at 50°C for 48 hours.
  • the solubility of the active agent is increased thanks to the prepared solid dispersion, the solubility of the active agent by itself and in solid dispersion form in pH 6.8 phosphate buffer is determined.
  • the amount of dissolved active agent is determined by identifying the active agent in the solutions inside the buffer mixed for 24 hours in the horizontal agitator.
  • the solubility of atorvastatin in solid dispersions prepared at the ratios of 1:9 and 1: 12 (active agent: polymer) increases.
  • thermograms of the drug and the polymer are taken by themselves and these results are compared with the endothermic peaks of the drug:polymer solid dispersion and physical mixtures.
  • XRD study the characteristic diffraction peaks reflecting the high crystalline properties of atorvastatin are compared with the peaks of the drug:polymer solid dispersion and physical mixtures.
  • Figures 5-7 show the thermograms of the DSC study according to the polymers.
  • thermograms it can be seen that the endothermic peak (165°C) of atorvastatin disappears in physical mixtures and solid dispersions. This shows that the crystalline properties of the drugs have transformed into an amorphous structure.
  • XRD X-ray diffraction
  • bioadhesion values of the gel compositions which are prepared based on the solid dispersions prepared with Atorvastatin and PEG 6000, Pluronic F-68, base chitosan polymers, are shown in Table 5. Table 5. Bioadhesive properties of gel formulations.
  • bioadhesion values of the compositions of the prepared solid dispersions inside chitosan gels are higher than the bioadhesion values of atorvastatin by itself in chitosan gels (gK-AT and gP-AT).
  • the increase of the adhesive property increases the duration for which the drug stays in the area of administration.
  • PEG 6000-atorvastatin solid dispersions with the melting method; first of all, PEG 6000 is heated in the water bath at 58°C until it melts, then atorvastatin is added onto this melt and it is constantly mixed until a homogeneous dispersion is obtained. Dispersions are kept in room temperature until solid and then, they are pulverized inside the porcelain mortar. Gel composition is prepared with base chitosan in 1% diluted lactic acid solution. The solid dispersion which was pulverized in the mortar before is added to this gel.
  • TNF-a pro-inflammatory cytokines
  • IL-6 interleukin
  • IL-8 interleukin 8
  • hGF human gingival fibroblast
  • the interleukin concentrations released from the cells with Human IL-8, IL- 1 beta, IL-6 and IL-10 Elisa analysis kits are measured and it is checked whether the interleukins (cytokines) are inhibited or not.
  • the cytokine levels measured after the administration of the compositions are compared with the cytokine levels of the control (HGF TNF (+)). Observing low cytokine levels show that there is high anti-inflammatory effect.
  • Chitosan gels containing solid dispersions at the ratio of 1: 12 (active agent: polymer), where the solubility of atorvastatin increases more, have been administered to the cells stimulated for determining the in vitro anti-inflammatory effect in this study. Examination of the I n Vivo Anti- inflammatorv Effect :
  • gel compositions containing 2% (w/v) atorvastatin have been administered to rats which developed periodontal disease and these groups are compared with the control groups to evaluate the effects on the treatment of the periodontal disease.
  • a periodontitis model induced with ligatures has been developed in the rats.
  • ligatures have been tied to the upper 2nd molar teeth of the rats and at the end of Day 11, it has been observed that the disease has developed with the presence of bleeding red gums [6], the ligatures have been removed and gel administration has started.
  • ligatures are removed and the rats are divided into experiment groups. Periodontal disease is developed in the groups other than the healthy control group. Gels containing 2 mg atorvastatin are locally administered every other day until the end of the study to the gingiva of rats which have periodontal pockets. Rats from each group are sacrificed at the ends of Week 1, 2, 3 and 4; and the jawbones are removed for histopathological and microbiological examination. Sections are taken from the removed jawbones, dyed with hematoxylin and eosin and the histopathological examination of each section is performed under a light microscope.
  • the gingiva surrounding the jaws of rats from each group is removed with scalpel to be placed inside distilled water containing 0.2 mL protease inhibitor cocktail and is stored at -80°C in refrigerators for microbiological examination. Then the tissues are thawed and the homogenized and released IL-6, IL-8, IL-10 and IL-lbeta levels are measured with Rat ELISA kits. Anti-inflammatory effects of the formulations are evaluated by checking the cytokine levels. Observing low cytokine levels show that there is high anti-inflammatory effect.
  • experiment groups gK-ML-dPEG12-AT which has exhibited high anti-inflammatory properties in, in vitro studies is selected among the gel formulations containing solid dispersions.
  • the experiment groups are as follows: Table 6. Experiment groups
  • Figures 11-14 show the IL-6, IL-8, IL-10 and IL-lbeta levels measured after the administration of the formulations in cell culture medium by using in vitro human gingival fibroblast cells (hGF). All the prepared 2% (w/v) gel compositions exhibit higher anti-inflammatory effect compared to the control. Observing low cytokine levels show that there is a high anti-inflammatory property.
  • Figure 11 shows the levels of IL-6 which is an inflammation indicator.
  • the compositions of the prepared atorvastatin solid dispersions in chitosan gel exhibit high anti-inflammatory effect by reducing the IL-6 levels more compared to atorvastatin administered by itself (AT) and chitosan gels administered by themselves (gK and gP).
  • compositions of the atorvastatin solid dispersions prepared by using PEG 6000 with various methods and the compositions of solid dispersions inside chitosan gels prepared by using Pluronic F-68 with the co-evaporation method compared to composition of atorvastatin by itself in chitosan gel (gK-AT and gP-AT).
  • the anti-inflammatory effect achieved after atorvastatin is added to the chitosan gel by increasing its solubility via the solid dispersion method is surprisingly higher than atorvastatin added to chitosan gel without increasing its solubility.
  • Figure 12 shows the levels of IL-8 which is another inflammation indicator.
  • Lower IL-8 levels are observed with the compositions of atorvastatin, the solubility of which is increased via the solid dispersion method, inside chitosan gels compared to chitosan gels by themselves (gK and gP) and atorvastatin by itself (AT); so, the anti-inflammatory effects of the former are higher.
  • the anti-inflammatory effects are surprisingly higher for these chitosan gels containing the solid dispersion form of atorvastatin, compared to atorvastatin added inside chitosan gel without increasing its solubility.
  • Figures 13 and 14 show that higher anti-inflammatory effect is achieved with all gel compositions compared to the control and that the anti-inflammatory effect increases with the chitosan gel compositions containing the solid dispersion forms of atorvastatin.
  • cytokine amounts are determined from the gingival tissue of the jaws of the rats sacrificed at the ends of week 1, 2, 3 and 4 by using the Rat ELISA kits.
  • Figures 15-18 show the IL-6, IL-8, IL-10 and IL-lbeta levels in the gingival tissues of the rats in each week. The obtained cytokine levels are used for evaluating whether the administered formulations have an anti-inflammatory effect or not. Observing low cytokine levels show that there is high anti-inflammatory effect. Higher anti-inflammatory effect is obtained with all the prepared gel formulations compared to the control group with periodontitis ((+) control).
  • both the base chitosan gel containing the solid dispersion form of atorvastatin and the base and water soluble chitosan gels containing only atorvastatin suppress cytokines and have higher anti-inflammatory effect compared to the (+) control group.

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Abstract

The present invention relates to a pharmaceutical composition containing atorvastatin, which is an active agent belonging to the statin group which has an anti-inflammatory effect, and/or a pharmaceutically acceptable salt thereof and at least one carrier polymer; and is developed in order to be used in the treatment of periodontal diseases (periodontitis). Thanks to this locally administered composition; the active agent with improved solubility is combined with the chitosan gels used as the carrier system to obtain high anti-inflammatory effect.

Description

A PHARMACEUTI CAL COMPOSI Tl ON CONTAI Nl NG ATORVASTATI N
Technical Field of the I nvention
The present invention relates to a pharmaceutical composition containing atorvastatin, which is an active agent belonging to the statin group which has an anti-inflammatory effect, and/or a pharmaceutically acceptable salt thereof and at least one carrier polymer; and is developed in order to be used in the treatment of periodontal diseases. Thanks to this locally administered composition; the active agent with improved solubility is combined with the chitosan gels used as the carrier system to obtain high anti-inflammatory effect.
Known State of the Art (Prior Art)
Periodontal diseases are inflammatory diseases which affect the gingiva and the other tissues supporting the teeth. Periodontal diseases start with gingivitis. Periodontitis is a more advanced stage of periodontal diseases. Alveolar bone is damaged in addition to the other tissues supporting the teeth. "Periodontal pockets" occur between the teeth and the gingiva. The presence of periodontal pockets facilitates the settling of the infection and the progression of the diseases. As the diseases progresses, the teeth start to become loose and this may lead to extraction. Thinning of the alveolar bone, degeneration of the collagen fibers, loss of periodontal ligaments and loss of attachment are the other outcomes of the disease.
The first treatment in periodontal diseases is the scaling of bacterial plaque and tartar which cause the disease to progress. For this purpose; mechanical practices such as tooth surface cleaning and root planning are performed. However; these practices are limited to only being the first line treatment in periodontal diseases and the bacteria which cannot be removed or accessed due to their attachment to the periodontal tissues cause the disease to progress and lead to loss of attachment and deterioration of the alveolar bone and the periodontal ligament.
In the known art; antibiotics such as tetracycline, metronidazole, clindamycin, amoxicillin, penicillin and cephalosporin are administered in systemic manner for treatment. Feres et al. mention the systemic use of antibiotics in their article for the treatment of periodontal disease {Systemic antibiotics in the treatment of periodontitis, Periodontology 2000, 67(1), 131-186, 2015). However, as the use of antibiotics cause severe bacterial resistance, the use of antibiotics to which the pathogens are resistant fails in stopping the progression of the disease. Furthermore, the use of systemic drugs for treatment causes the drug to suffer the hepatic first pass effect, to undergo enzymatic or hydrolytic decomposition in the gastrointestinal tract and consequently to have a lower amount of drug that reaches the area of effect. Thus, drugs with higher dose are used in this practice, which is undesirable for human health.
Topical chemical practices also exist in addition to the mechanical cleaning performed to control and prevent the formation of plaque. Mouthwash solutions are frequently used in the topical practice. For this purpose, the most number of studies are related to chlorhexidine which an antimicrobial agent is proven to inhibit dental plaque. However, these solutions have certain disadvantages such as the necessity to use them several times a day, and the inability of the solution to stay at the area of effect for long period of time due to the lack of adhesion to the periodontal pocket occurring in periodontal disease and disposal by saliva.
Local drug treatment outclasses systemic drug treatment thanks to advantages such as high drug concentration at the area of administration, low side effects, non-necessity of daily administration. However; due to the aforementioned disadvantages, bioadhesive drug formulations are required for preventing the removal of the locally administered drug from the area of administration due to the presence of saliva and tongue movement, and for ensuring that said drug stays at the area long enough to create the desired effect.
Statins are drugs which are used to treat hypercholesterolemia in atherosclerosis and cardiovascular system disorders, and they regulate the cholesterol level in blood by inhibiting the biosynthesis of cholesterol in the liver. They reduce the levels of low density lipoprotein (LDL) in the plasma by acting as 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG CoA reductase) inhibitors. For this purpose; various drugs belonging to the statin group such as atorvastatin, simvastatin, rosuvastatin, lovastatin, fluvastatin and pravastatin.
Recent studies found that statins exhibit pleiotropic effects. "Pleiotropic effect" is the effect exhibited by the drug apart from the effect it is specifically developed for. Besides their lipid lowering effects, statins have anabolic effect and anti-inflammatory effect on bones as their pleiotropic effects. In the known art, there are certain studies showing that the use of statin reduces the risk of inflammation [1, 2]. Patent document no: US 2001006656 Al, describes an aerosol formulation which contains HMG- CoA reductase inhibitor and which is used for spray administration. Said HMG-CoA reductase inhibitor can be lovastatin, pravastatin, simvastatin, cerivastatin, fluvastatin, atorvastatin or mevastatin. The invention is developed in order to be used for the treatment of respiratory system diseases related to inflammation such as asthma, pneumonia and chronic bronchitis.
In the inflammation process, IL-6 (interleukin-6), IL-8 (interleukin-8), IL-10 (interleukin-10), IL- lbeta (interleukin-lbeta) cytokines, which are inflammation indicators, play an important role. It is known that the cytokine levels, which are inflammation indicators, are decreased thanks to the use of statin [3].
Atorvastatin; is a drug with anti-inflammatory effects which belongs to the statin group and has low water solubility. It does not dissolve in aqueous solutions at and below pH 4; and it slightly dissolves in distilled water and pH 7.4 phosphate buffer. The chemical formula of Atorvastatin, the chemical name of which is [(3R,5R)-7-[2-(4-fluorophenyl)-3-phenyl-4-(phenylcarbamoyl)-5- propane-2-ylpyrol-l-yl]-3,5-dihydroxyheptanoic acid], is shown below:
Figure imgf000005_0001
Methods for increasing solubility or methods for increasing the surface area for solution are physical and chemical modifications. Physical modifications are processes such as decreasing particle sizes via micronisation, changing crystal properties, creating polymorphs, creating complex with cyclodextrin, solutionizing, preparing the eutectic mixture of the drug inside the carrier and preparing the solid dispersion thereof. Chemical modifications are processes such as creating a soluble prodrug and creating the salts of the drug. Chitosan is a natural polymer obtained by the partial deacetylation chitin which is present in the shells of shellfish such as crabs and shrimps, shells of insects and cell walls of fungi. Physical- chemical and bioactive properties of chitosan, which is a cationic and biodegradable polymer, change depending on properties such as the level of deacetylation and molecular weight. In addition to chitosan being a biocompatible, biodegradable, non-toxic and non-allergenic natural polymer; it has antimicrobial, hemostatic, wound healing and tissue regenerating biological effects. Moreover, since it is a bioadhesive polymer, it is typically preferred in carrier systems which are locally administered [4]. The structure of chitosan, which is a copolymer of D- glucosamine and N-acetyl-D-glucosamine, is presented below.
Figure imgf000006_0001
Formula-2
In the technical field of the invention, innovation regarding compositions having antiinflammatory effects is required. In the known art, the use of atorvastatin as an antiinflammatory drug in the periodontal disease is known. However, there is no system where the solubility of atorvastatin of the invention is increased and atorvastatin is used inside a chitosan based gel in order to be administered locally in the treatment of the periodontal disease.
Thanks to the present invention; the solubility problem of atorvastatin in locally administered pharmaceutical compositions developed for the treatment of the periodontal disease is eliminated. Furthermore, higher effects can be achieved with lower doses thanks to the local administration of the invention, and the drug is not easily removed by the presence of saliva and/or tongue movements, thereby enabling the drug to have an effect on the area of administration for a long period of time. Thus, bioavailability increases, the frequency of side effects decreases and patient compatibility rises. Also, there are various advantages such as ease of administration and prolonged presence at the area of administration due to the bioadhesive properties of the drug.
Brief Description and Objective of the I nvention
The present invention relates to a new pharmaceutical composition developed by eliminating the solubility problem of atorvastatin, which belongs to the statin group, through various pharmaceutical processes in order to be used in the treatment of periodontal diseases.
Higher effects at lower doses can be achieved through local administration thanks to the present invention. Thus, bioavailability and patient compatibility increases, whereas the frequency of side effects decreases. In addition to the developed gel being easy to administer, it is not easily removed from the area of administration by the presence of saliva and/or tongue movements and thereby the drug has a prolonged effect at said area of administration. Moreover, as the used chitosan with bioadhesive properties is biocompatible and has its own anti-inflammatory effect, it is preferred as the carrier system. Therefore, the anti-inflammatory effects of the formulations prepared with chitosan gels are increased.
In addition to these, local drug treatment outclasses systemic drug treatment thanks to advantages such as high drug concentration, low side effects, non-necessity of daily administration.
Description of the Figures Explaining the I nvention
Figure 1 . Release graph for active substance in gel formulations
Figure 2. Release graph for active substance in gels prepared with PEG 6000 solid dispersions
Figure 3. Release graph for active substance in gels prepared with Pluronic F-68 solid dispersions
Figure 4. Release graph for active substance in gels prepared with base chitosan solid dispersions
Figure 5. DSC thermograms of Atorvastatin and PEG 6000
Figure 6. DSC thermograms of Atorvastatin and Pluronic F-68 Figure 7. DSC thermograms of Atorvastatin and base chitosan
Figure 8. X-ray diffraction of Atorvastatin and PEG 6000
Figure 9. X-ray diffraction of Atorvastatin and Pluronic F-68
Figure 10. X-ray diffraction of Atorvastatin and base chitosan
Figure 1 1 . IL-6 levels 4 hours after administration in hGF cell culture in (1/2) dilution of formulations
Figure 12. IL-8 levels 4 hours after administration in hGF cell culture in (1/2) dilution of formulations
Figure 13. IL-10 levels 4 hours after administration in hGF cell culture in (1/2) dilution of formulations
Figure 14. IL-lbeta levels 4 hours after administration in hGF cell culture in (1/2) dilution of formulations
Figure 15. IL-6 levels measured from the gingival tissues
Figure 16. IL-8 levels measured from the gingival tissues
FFiigguurree 1177.. IL-10 levels measured from the gingival tissues
Figure 18. IL-lbeta levels measured from the gingival tissues
Figure 19. Histological micrograph of the periodontal tissues of the healthy control group ((-) control) (at the end of Week 1)
Figure 20. Histological micrograph of the periodontal tissues of the control group with periodontal disease without administration ((+) control) (at the end of Week 1)
Figure 21 . Histological micrograph of the periodontal tissues of the group administered with Atorvastatin (AT) (at the end of Week 1)
Figure 22. Histological micrograph of the periodontal tissues of the group administered with base chitosan gel containing Atorvastatin (gK-AT) (at the end of Week 1)
Figure 23. Histological micrograph of the periodontal tissues of the group administered with water soluble chitosan gel containing Atorvastatin (gP-AT) (at the end of Week 1)
Figure 24. Atorvastatin prepared by the melting method: Histological micrograph of the periodontal tissues of the group administered with base chitosan gel containing PEG 6000 (1: 12) solid dispersion (gK~ML-dPEG12 - AT) (at the end of Week 1) Detailed Description of the I nvention
The present invention relates to a gel, fiber, sponge, film, micro or nanoparticle system administered locally with an active agent selected from the statin group and/or a pharmaceutically acceptable salt thereof and at least one carrier polymer for use in the treatment of the periodontal disease, and the production method of said pharmaceutical composition. The active agent is selected among atorvastatin, simvastatin, rosuvastatin, lovastatin, fluvastatin, pravastatin from the statin group and/or the acceptable salts thereof. The preferred form of the invention is a gel composition of atorvastatin and/or and acceptable salt thereof. Atorvastatin which is used in the gel composition is selected within the range of 0.5-4% (weight/volume, w/v). Atorvastatin and/or an acceptable salt thereof are used preferably as 2% (w/v) in the gel composition. Atorvastatin, which is known to exhibit properties regulating the cholesterol level and reducing lipids when used in high doses, exhibits an anti-inflammatory effect when used in low doses in our study. Furthermore, as the drugs used in high doses damage the kidneys and cause renal toxicity, it is important to choose a treatment dosage where the drug exhibits an effect in low doses.
The carrier polymer used in the preparation of the gel composition is selected among polymers with bioadhesive properties, such as chitosans which have various molecular weights (MW), various deacetylation levels and various solubility, sodium alginate, hyraluronic acid, pectin, agarose, gelatin, carrageenan, aminodextran, hydroxypropyl cellulose (HPC), polyethylene glycol (PEG), polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP), ethyl cellulose, hydroxy ethyl cellulose, polycarbophil, methylvinyl ether and methacrylic acid copolymer, polyacrylates, polyalkylcyanoacrylate, acrylic acid and ehtlyhexylacyrlate copolymer, polymethacrylate, carbopol, methyl hydroxy ethyl cellulose, methyl cellulose, hydroxy propyl methyl cellulose (HPMC) and carboxy methyl cellulose (CMC).
In the preferred form of the present invention, atorvastatin with increased solubility is used and as the carrier polymer, base chitosan (dissolves in diluted acidic solutions) which is a biodegradable, biocompatible, natural, non-toxic polymer or water soluble chitosan containing different salts thereof, is used. Chitosans are preferred in the formulation due to the antimicrobial, anti-inflammatory, tissue regenerating, wound and bone healing etc. bioactive properties they possess. Also, the physical-chemical and bioactive properties of chitosans with various molecular weights (MW), various deacetylation levels and various solubility vary. Since the bioactive properties they possess can be varied depending on the selected type of chitosan, it is important to show that the chitosan used in the formulation has a high anti-inflammatory effect. The anti-inflammatory activity of the drug increases thanks to the base chitosan and water soluble chitosan used in the present invention. In the invention, preferably, chitosan with 85% deacetylation level is used as base chitosan and chloride salt of chitosan with 86% deacetylation level is used as water soluble chitosan. Chitosan can be used in the range of 0.1% - 10% (w/v) in the formulation. The chitosan used in the invention is at the rate of 2% (w/v).
In the administration of the formulation to be used in the local treatment of the disease via dose adjusted systems (applicator) where the drug is injected without a needle, the properties of injectability and viscosity need to be appropriate for the injection of the drug in said area. Since viscosity increases as the chitosan dose used in the preparation of the gel increases, the property of injectability of the obtained viscous gel decreases and it becomes harder to administer the drug in the desired dosage. On the other hand, since viscosity will be low in the chitosan gel formulation used in lower doses and a highly fluid gel will be obtained, the injectability of the drug will decrease; furthermore, since the bioadhesive properties of the chitosan gel used in lower doses will also be low, it will become harder for the drug to adhere to the area of administration and the drug will not be able to stay in said area for a long period of time. The properties of injectability and bioadhesion of the gels used in the invention with 2% chitosan concentration are quite high.
Base chitosan dissolves in diluted acidic solutions. For this purpose; organic acids such as acetic acid, lactic acid, citric acid, malic acid and formic acid are used in the invention. These acid solutions can be prepared as 0.1% - 5%. In the invention, preferably, 1% diluted lactic acid solution is prepared by using diluted lactic acid. As the percentage of the lactic acid used in obtaining the composition changes, the properties of injectability with syringe, viscosity and bioadhesion of the gel also change. The gel formulation to be prepared with 2% chitosan which exhibits properties of pseudoplastic flow and high bioadhesion, can be obtained by preparing the lactic acid solution as 1%.
Primarily, release studies have been performed with only atorvastatin, and the mixture thereof with base chitosan and water soluble chitosan gels. Release studies are performed at 37°C by using Franz diffusion cells with a 2.01 cm2 diffusion area and 20 mL receptor volume. pH 6.8 phosphate buffer is used as the diffusion medium. 1 mL samples are collected at certain time intervals and the same amount of buffer is added instead. The amount of active agent in the collected samples is measured spectrophotometrically. Figure 1 shows the graph for the release studies. Atorvastatin is dissolved in PEG400. Release is found to be low when the active agent concentrations released from the gels are compared to this PEG400-AT mixture. Solubility of atorvastatin needs to be increased in order to improve the release profile. In the invention, the method of creating solid dispersion is utilized in order to increase the solubility of atorvastatin. The method of creating solid dispersion is a more practical approach for increasing solubility as it is easier and more feasible than chemical modifications. Thanks to the method of creating solid dispersion, particle size is reduced, wettability and porosity of the particles increase, drugs with crystal structures which have low water solubility are transformed into amorphous form and thus, solubility increases. Also, the bioavailability of the drugs increases thanks to the method of creating solid dispersion. Methods of melting, hot melt extrusion, co-evaporation, rotating evaporator, spray drying, freeze drying, spray freeze drying, supercritical antisolvent, co-precipitation, electrostatic spinning, co-milling and kneading are used in order to create solid dispersions.
In the invention, melting, co-evaporation, co-milling and kneading are used as the methods for creating solid dispersions. At high temperatures, degradation of the drug and the polymer may occur. Moreover, loss of drug may occur due to the drug and polymer sticking to the walls of the tools used in creating solid dispersions. Due to these disadvantages; methods of co-evaporation, melting, co-milling and kneading, which do not require heat or which operate at low temperatures, are preferred and no drug loss occurs. Additionally, these methods are preferred in the invention as it is easier to prepare solid dispersions with these methods compared to others.
The polymers used in creating solid dispersions can be selected among: cellulose derivatives such as hydroxypropyl methyl cellulose (HPMC), ethyl cellulose, hydroxypropyl cellulose (HPC), sodium carboxymethyl cellulose (NaCMC), starch derivatives, polyvinyl pyrrolidone, eudragit, polyethylene glycols, polymethacrylates, chitosans with various deacetylation, various molecular weights and various solubility, inulin, compritol 888 ATO, gelucire, pluronics, gelatin, cyclodextrins, Kollidon, Avicel 200, Synperonic and Plasdone. The water solubility of the drug with crystal structure is increased via the method of creating solid dispersions. Polyethylene glycol, pluronic and chitosan polymers are used in the invention for creating solid dispersions. In the preferred form of the invention, polyethylene glycol 6000 (PEG 6000), pluronic F-68 (PL F- 68) and base chitosan polymers are used. The reason for using polyethylene glycol 6000 and pluronic F-68 among the water soluble polymers is that it is more effective to utilize polymers with high water solubility in order to increase the water solubility of the drug. The reason for using chitosan is to ensure that the anti-inflammatory properties of the drug are further increased by making use of the bioactive properties of chitosan. As solid dispersions cannot be created with water soluble chitosan, it is not included among the polymers selected for creating solid dispersions.
The polymers used in preparing solid dispersions can be used at the ratios of 1:0.1 - 1:25 (active agent: polymer). The obtained values of solubility, release and dissolution change depending on the various polymers, various active agent: polymer ratios, various methods and various active agents used in the solid dispersions. For this reason, it is not possible to generalize by saying that a solid dispersion with higher solubility, release and dissolution values is going to be obtained due to increased or decreased polymer ratios. For example, solid dispersions of piroxicam with PVP K-30 have been prepared at the active agent: polymer ratios of 1:0.5, 1: 1, 1:2, 1:3, 1:4, 1:5 and 1:6; and the highest dissolution value has been obtained with the solid dispersion prepared at the ratio of 1:4 [5]. In the invention, solid dispersions of atorvastatin with the ratios of 1: 1, 1:3, 1:9 and 1: 12 (active agent: polymer) have been prepared. These prepared solid dispersions have been mixed with chitosan gels and the release studies of these compositions have been performed. As a result of these studies; the active agent release of the chitosan gel compositions containing solid dispersions prepared at ratios of 1: 1 and 1:3 (active agent: polymer) are lower than the gel compositions of the solid dispersions prepared at ratios of 1:9 and 1: 12 (active agent: polymer). The developed gel compositions are shown in Table 1. Table 1 . Gel formulations developed according to the invention
Figure imgf000012_0001
gCG-dK12 Base chitosan (1: 12) (co-milling) + 1% lactic acid solution gP-CE-dPL12 PEG 6000 (1: 12) (co-evaporation) + water soluble chitosan
gel
gP-CE-dPEG12 Pluronic F-68 (1: 12) (co-evaporation) + water soluble
chitosan gel
gP-ML-dPL12 PEG 6000 (1: 12) (melting) + water soluble chitosan gel
gP-ML-dPEG12 Pluronic F-68 (1: 12) (melting) + water soluble chitosan gel
Drug Loaded
gK-AT Atorvastatin+Base chitosan gel
gP-AT Atorvastatin+Water soluble chitosan gel
PEG 400-AT Atorvastatin+PEG 6000
gK-ML-dPEG3-AT AT: PEG 6000 (1:3) (melting)+base chitosan gel
gK-ML-dPL3-AT AT:Pluronic F-68 (1:3) (melting)+base chitosan gel
gK-CE-dKl-AT AT: Base chitosan (1: 1) (solvent)+base chitosan gel
gP-ML-dPEG3-AT AT: PEG 6000 (1:3) (melting)+water soluble chitosan gel
gP-ML-dPL3-AT AT:Pluronic F-68 (1:3) (melting)+water soluble chitosan gel gK-ML-dPEG9-AT AT: PEG 6000 (1:9) (melting) + base chitosan gel
gK-ML-dPL9-AT AT:Pluronic F-68 (1:9) (melting) + base chitosan gel
gK-CE-dPEG9-AT AT: PEG 6000 (1:9) (co-evaporation) + base chitosan gel
gK-CE-dPL9-AT AT:Pluronic F-68 (1:9) (co-evaporation) + base chitosan gel
gKN-dK9-AT AT: Base chitosan (1:9) (kneading) + 1% lactic acid solution
gCG-dK9-AT AT:Base chitosan (1:9) (co-milling) + 1% lactic acid solution
gK-ML-dPEG12-AT AT: PEG 6000 (1: 12) (melting method) + base chitosan gel
gK-ML-dPL12-AT AT:Pluronic F-68 (1: 12) (melting method) + base chitosan gel gK-CE-dPEG12-AT AT: PEG 6000 (1: 12) (co-evaporation) + base chitosan gel
gK-CE-dPL12-AT AT:Pluronic F-68 (1: 12) (co-evaporation) + base chitosan gel gKN-dK12-AT AT:Base chitosan (1: 12) (kneading) + 1% lactic acid solution gCG-dK12-AT AT: Base chitosan (1: 12) (co-milling) + 1% lactic acid solution gP-CE-dPL12-AT AT: PEG 6000 (1: 12) (co-evaporation) + water soluble
chitosan gel
gP-CE-dPEG12-AT AT:Pluronic F-68 (1: 12) (co-evaporation) + water soluble
chitosan gel
gP-ML-dPL12-AT AT: PEG 6000 (1: 12) (melting) + water soluble chitosan gel
gP-ML-dPEG12-AT AT:Pluronic F-68 (1: 12) (melting) + water soluble chitosan gel
Figures 2-4 show the active agent release graphs of chitosan gels containing atorvastatin solid dispersions. Release graphs are presented separately according to the type of the used polymer. It can be seen that the active agent release from the chitosan gel compositions containing solid dispersions prepared at ratios of 1: 1 and 1:3 (active agent: polymer) are low.
On the other hand, the active agent release from the chitosan gel compositions containing solid dispersions prepared at ratios of 1:9 and 1: 12 (active agent: polymer) are higher than the release of atorvastatin from base and water soluble chitosan gels (gK-AT and gP-AT) in Figure 1. For this reason; ratios of 1:9 and 1: 12 (active agent: polymer) are preferred over the ratios of 1: 1 and 1:3 (active agent: polymer) in the invention. The table below shows the solid dispersions prepared at the active agent:polymer ratios of 1:9 and 1: 12 with polymers and various preparation methods.
Table 2. Preparation of the solid dispersion formulation
Figure imgf000014_0001
Drug release from the gels of the solid dispersions prepared with base chitosan decreases when the methods of melting and co-evaporation are used, and said release from gels increases when the methods of co-milling and kneading are used. No increase in the drug solubility is observed when the methods of co-milling and kneading are used with PEG 6000. On the other hand, solid dispersions cannot be created due to the properties of the polymer when the methods of co- milling and kneading are used with Pluronic F-68. For this reason; the methods of melting and co-evaporation for PEG 6000 and Pluronic F-68 and the methods of co-milling and kneading for base chitosan are preferred in the invention for having the highest solubility and release values.
In order to prepare PEG 6000-atorvastatin and Pluronic F-68-atorva statin solid dispersions with the melting method; first of all, the polymer is heated in the water bath at 40-100°C until it melts, then atorvastatin is added onto this melt and it is constantly mixed until a homogeneous dispersion is obtained. In the invention, preferably, 58°C is selected as the temperature where the polymers melt in order for the drug to not degrade, atorvastatin is added to the molten polymer and mixing is performed. Mixtures prepared with Pluronic F-68 and PEG 6000 are kept until solid. Preferably, they are kept in the refrigerator or the desiccator. Afterwards, both mixtures are pulverized separately.
In order to prepare PEG 6000-atorvastatin and Pluronic F-68-atorva statin solid dispersions with the co-evaporation method; atorvastatin and the polymer are dissolved in an organic solvent, preferably methanol, and afterwards the solvent is evaporated. Methanol is chosen as the solvent because atorvastatin easily dissolved in methanol. However, since methanol causes the experiment to yield incorrect results by causing cells to die in the cell culture studies carried out to show the anti-inflammatory effects of the composition; there should be no traces of methanol in the prepared composition. Therefore, methanol is preferably evaporated in the rotating evaporator at 30°C-80°C. In the invention, preferably, 40°C, which is the temperature at which the drug will not degrade and the minimum temperature where the organic solvent (methanol) evaporates, is used and the organic solvent is completely evaporated. The obtained PEG 6000- atorvastatin and Pluronic F-68-atorvastatin mixtures are dried until no moisture remains. Preferably, they are kept inside the desiccator or the refrigerator for 24 hours. Afterwards, both mixtures are pulverized separately.
In order to prepare atorvastatinxhitosan solid dispersion with the co-milling method; physical atorvastatin and polymer mixtures with the ratios of 1:9 and 1: 12 are mixed inside the ball mill and pulverized. The mixing process is carried out for 1-24 hours at 100-600 rpm. In the invention, the powder mixture is preferably prepared by mixing for 1 hour at 300 rpm. The reason for choosing these conditions is that mixture sticks to the inner walls or the balls of the device when mixed at a higher speed or for a longer time. Therefore, a speed is selected where the highest yield is achieved in the shortest time.
In order to prepare atorvastatin: chitosan solid dispersion with the kneading method; physical atorvastatin and polymer mixtures with the ratios of 1:9 and 1: 12 are placed inside the porcelain mortar and are mixed with organic solvent or organic solvent: water mixture, the amount of which is 1 to 10 times amount the weighed powder. The organic solvent: water mixture to be used is selected from the range of 0.1: 1 - 20: 1. The obtained mixture is dried inside a dryer at a temperature selected between 10°C - 80°C by keeping said mixture therein for 1 - 5 days. In the present invention, the mixture of atorvastatin: polymer is mixed by wetting it with a methanokwater (6: 1) mixture 1.5 times the amount of the weighed powder. Since the water solubility of atorvastatin is quite low and it can easily dissolve in methanol, it is important to obtain a solution mixture prepared at the correct ratios. Furthermore, since methanol may cause the experiment to not yield the correct results by causing cells to die in the cell culture studies carried out to show the anti-inflammatory effects of the composition; there should be no traces of methanol in the prepared composition. In other words, the methanol to be used should be present in an amount that will allow atorvastatin to dissolve and it should be used minimally in a way that it does not remain in the formulation after drying. For this reason, the obtained mixture is kept inside the oven at 50°C for 48 hours.
Table 3. Prepared solid dispersions
Figure imgf000016_0001
Examination of the solubility of solid dispersion:
In order to see whether the solubility of the active agent is increased thanks to the prepared solid dispersion, the solubility of the active agent by itself and in solid dispersion form in pH 6.8 phosphate buffer is determined. The amount of dissolved active agent is determined by identifying the active agent in the solutions inside the buffer mixed for 24 hours in the horizontal agitator. In the study, it is observed that the solubility of atorvastatin in solid dispersions prepared at the ratios of 1:9 and 1: 12 (active agent: polymer) increases.
Table 4. Solubility of the atorvastatin solid dispersions
Figure imgf000017_0001
The solubility of atorvastatin in pH 6.8 phosphate buffer is quite low; whereas its solubility significantly increases in solid dispersion form. The highest solubility value is obtained with the solid dispersion prepared with the co-evaporation method and pluronic F-68, where the solubility of atorvastatin increases approximately 35 times.
The loss of the drug's crystal properties and transformation into amorphous form is supported by the characterization studies performed with drug-polymer dispersion, the differential scanning calorimetry (DSC) studies and the X-ray diffraction (XRD) studies (Figures 5-10). In the DSC study, thermograms of the drug and the polymer are taken by themselves and these results are compared with the endothermic peaks of the drug:polymer solid dispersion and physical mixtures. In the XRD study, the characteristic diffraction peaks reflecting the high crystalline properties of atorvastatin are compared with the peaks of the drug:polymer solid dispersion and physical mixtures. Figures 5-7 show the thermograms of the DSC study according to the polymers. In the thermograms, it can be seen that the endothermic peak (165°C) of atorvastatin disappears in physical mixtures and solid dispersions. This shows that the crystalline properties of the drugs have transformed into an amorphous structure. In the X-ray diffraction (XRD) study presented in Figures 8-10, it can be observed that the intensive peaks characteristic to the crystalline structure of atorvastatin have disappeared in physical mixtures and solid dispersions. This shows, in line with the DSC study, that atorvastatin has transformed from crystal form to amorphous form. The bioadhesion values of the gel compositions which are prepared based on the solid dispersions prepared with Atorvastatin and PEG 6000, Pluronic F-68, base chitosan polymers, are shown in Table 5. Table 5. Bioadhesive properties of gel formulations.
Figure imgf000018_0001
The bioadhesion values of the compositions of the prepared solid dispersions inside chitosan gels are higher than the bioadhesion values of atorvastatin by itself in chitosan gels (gK-AT and gP-AT). The increase of the adhesive property increases the duration for which the drug stays in the area of administration. Example:
Figure imgf000019_0001
In order to prepare PEG 6000-atorvastatin solid dispersions with the melting method; first of all, PEG 6000 is heated in the water bath at 58°C until it melts, then atorvastatin is added onto this melt and it is constantly mixed until a homogeneous dispersion is obtained. Dispersions are kept in room temperature until solid and then, they are pulverized inside the porcelain mortar. Gel composition is prepared with base chitosan in 1% diluted lactic acid solution. The solid dispersion which was pulverized in the mortar before is added to this gel.
Determination of the I n Vitro Anti-inflammatorv Effect :
The formation of periodontal inflammation starts with the releasing of pro-inflammatory cytokines such as TNF-a, (tumor necrosis factor), IL-6 (interleukin), IL-8 etc. and as inflammation progresses, the amounts of these inflammation indicators gradually increases. In the invention, human gingival fibroblast (hGF) cells are used for determining the in vitro antiinflammatory effect. Cells are stimulated with TNF-a after they completely cover the surface of the wells. Compositions are administered to the stimulated wells and the results obtained after 4 hours are examined. The interleukin concentrations released from the cells with Human IL-8, IL- 1 beta, IL-6 and IL-10 Elisa analysis kits are measured and it is checked whether the interleukins (cytokines) are inhibited or not. The cytokine levels measured after the administration of the compositions are compared with the cytokine levels of the control (HGF TNF (+)). Observing low cytokine levels show that there is high anti-inflammatory effect. Chitosan gels containing solid dispersions at the ratio of 1: 12 (active agent: polymer), where the solubility of atorvastatin increases more, have been administered to the cells stimulated for determining the in vitro anti-inflammatory effect in this study. Examination of the I n Vivo Anti- inflammatorv Effect :
The study protocol followed for the in vivo experiments has been approved by Hacettepe University Animal Experiments Local Ethics Committee, with decision no: 2014/16 and has been carried out in the Medical-Surgical Research Laboratory allocated for Experimental Animals in Hacettepe University.
In this study, gel compositions containing 2% (w/v) atorvastatin have been administered to rats which developed periodontal disease and these groups are compared with the control groups to evaluate the effects on the treatment of the periodontal disease. For this purpose, first of all, a periodontitis model induced with ligatures has been developed in the rats. On Day 1, ligatures have been tied to the upper 2nd molar teeth of the rats and at the end of Day 11, it has been observed that the disease has developed with the presence of bleeding red gums [6], the ligatures have been removed and gel administration has started.
Experiment Protocol :
At the end of Day 11, ligatures are removed and the rats are divided into experiment groups. Periodontal disease is developed in the groups other than the healthy control group. Gels containing 2 mg atorvastatin are locally administered every other day until the end of the study to the gingiva of rats which have periodontal pockets. Rats from each group are sacrificed at the ends of Week 1, 2, 3 and 4; and the jawbones are removed for histopathological and microbiological examination. Sections are taken from the removed jawbones, dyed with hematoxylin and eosin and the histopathological examination of each section is performed under a light microscope. Also, the gingiva surrounding the jaws of rats from each group is removed with scalpel to be placed inside distilled water containing 0.2 mL protease inhibitor cocktail and is stored at -80°C in refrigerators for microbiological examination. Then the tissues are thawed and the homogenized and released IL-6, IL-8, IL-10 and IL-lbeta levels are measured with Rat ELISA kits. Anti-inflammatory effects of the formulations are evaluated by checking the cytokine levels. Observing low cytokine levels show that there is high anti-inflammatory effect.
In one of the experiment groups, gK-ML-dPEG12-AT which has exhibited high anti-inflammatory properties in, in vitro studies is selected among the gel formulations containing solid dispersions. The experiment groups are as follows: Table 6. Experiment groups
Figure imgf000021_0001
Results
Examination of the I n Vitro Anti-inflammatory Effect :
Figures 11-14 show the IL-6, IL-8, IL-10 and IL-lbeta levels measured after the administration of the formulations in cell culture medium by using in vitro human gingival fibroblast cells (hGF). All the prepared 2% (w/v) gel compositions exhibit higher anti-inflammatory effect compared to the control. Observing low cytokine levels show that there is a high anti-inflammatory property.
Figure 11 shows the levels of IL-6 which is an inflammation indicator. The compositions of the prepared atorvastatin solid dispersions in chitosan gel exhibit high anti-inflammatory effect by reducing the IL-6 levels more compared to atorvastatin administered by itself (AT) and chitosan gels administered by themselves (gK and gP).
Moreover; higher anti-inflammatory effect is obtained with the compositions of the atorvastatin solid dispersions prepared by using PEG 6000 with various methods and the compositions of solid dispersions inside chitosan gels prepared by using Pluronic F-68 with the co-evaporation method, compared to composition of atorvastatin by itself in chitosan gel (gK-AT and gP-AT). So, the anti-inflammatory effect achieved after atorvastatin is added to the chitosan gel by increasing its solubility via the solid dispersion method is surprisingly higher than atorvastatin added to chitosan gel without increasing its solubility.
Figure 12 shows the levels of IL-8 which is another inflammation indicator. Lower IL-8 levels are observed with the compositions of atorvastatin, the solubility of which is increased via the solid dispersion method, inside chitosan gels compared to chitosan gels by themselves (gK and gP) and atorvastatin by itself (AT); so, the anti-inflammatory effects of the former are higher. The anti-inflammatory effects are surprisingly higher for these chitosan gels containing the solid dispersion form of atorvastatin, compared to atorvastatin added inside chitosan gel without increasing its solubility.
Figures 13 and 14 show that higher anti-inflammatory effect is achieved with all gel compositions compared to the control and that the anti-inflammatory effect increases with the chitosan gel compositions containing the solid dispersion forms of atorvastatin.
Examination of the I n Vivo Anti- inflammatory Effect :
In the microbiological and histological studies performed on rats, comparisons are made between the groups where gel administration is performed on rats which developed periodontal diseases, and the control group with periodontitis where no administration is performed ((+) control) and the healthy control group ((-) control).
In the microbiological studies, cytokine amounts are determined from the gingival tissue of the jaws of the rats sacrificed at the ends of week 1, 2, 3 and 4 by using the Rat ELISA kits. Figures 15-18 show the IL-6, IL-8, IL-10 and IL-lbeta levels in the gingival tissues of the rats in each week. The obtained cytokine levels are used for evaluating whether the administered formulations have an anti-inflammatory effect or not. Observing low cytokine levels show that there is high anti-inflammatory effect. Higher anti-inflammatory effect is obtained with all the prepared gel formulations compared to the control group with periodontitis ((+) control). It can be seen that both the base chitosan gel containing the solid dispersion form of atorvastatin and the base and water soluble chitosan gels containing only atorvastatin suppress cytokines and have higher anti-inflammatory effect compared to the (+) control group.
The images of the sections belonging to the jawbones removed at the end of Week 1 in the histological studies under light microscope are presented in Figures 19-24; and the micrographs show alveolar bones from the various administration groups. Periodontal disease is characterized with the thinning of the alveolar bone. In Figure 19, the alveolar bone of the healthy group can be seen as intact without thinning, whereas in Figure 20 of the control group with periodontitis, alveolar bone has thinned and became smaller (the arrow with dots inside and shown with E¾4N indicates bone thinning; the arrow with horizontal lines and shown with ssssss^ indicates new bone formation).
In Figure 21, there is loss in the alveolar bone of the group administered with only atorvastatin; whereas in Figures 22-24, alveolar bone can be seen as a whole with the new bone formation in groups administered with the atorvastatin compositions inside chitosan gels. The obtained formulations can be used for the treatment of periodontal disease, as well as for the treatment of dermal or mucosal inflammations for topical purposes.
References
1. Jerwood, S. and J. Cohen, "Unexpected antimicrobial effect of statins. " Journal of antimicrobial chemotherapy, 2008. 61(2): p. 362-364.
2. Weitz-Schmidt, G., "Statins as anti-inflammatory agents. " Trends in pharmacological sciences, 2002. 23(10): p. 482-487.
3. Menge, T., H.-P. Hartung, and O. Stuve, "Statins— a cure-all for the brain?' Nature Reviews Neuroscience, 2005. 6(4): p. 325-331.
4. Senel, S., Potential applications of chitosan in oral mucosal delivery. Journal of Drug Delivery Science and Technology, 2010. 20(1): p. 23-32.
5. Tantishaiyakul, V., N. Kaewnopparat, and S. Ingkatawornwong. "Properties of solid dispersions of piroxicam in polyvinylpyrrolidone K-30." International journal of pharmaceutics, 1996, 143.1: 59-66. Queiroz-Junior, C, et al., "Role of systemic and local administration of selective inhibitors of cyclo-oxygenase 1 and 2 in an experimental model of periodontal disease in rats. " Journal of periodontal research, 2009. 44(2): p. 153-160.

Claims

CLAI MS
The preparation method for a pharmaceutical gel composition containing atorvastatin and/or a pharmaceutically acceptable salt thereof as active agent and at least one carrier polymer; characterized in that it comprises the steps of:
- preparing solid dispersions containing active agent:polymer at the ratios between the range of 1:0.1 - 1:25 with methods for creating solid dispersions,
- preparing gel composition and
- adding the solid dispersions into the prepared gel composition.
A method according to Claim 1; characterized in that the active agent is atorvastatin calcium.
A method according to Claim 1; characterized in that the carrier polymers are bioadhesive polymers selected among the group containing chitosans which have various molecular weights (MW), various deacetylation levels and various solubility, sodium alginate, hyraluronic acid, pectin, agarose, gelatin, carrageenan, aminodextran, hydroxypropyl cellulose (HPC), polyethylene glycol (PEG), polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP), ethyl cellulose, hydroxy ethyl cellulose, polycarbophil, methylvinyl ether and methacrylic acid copolymer, polyacrylates, polyalkylcyanoacrylate, acrylic acid and ehtlyhexylacyrlate copolymer, polymethacrylate, carbopol, methyl hydroxy ethyl cellulose, methyl cellulose, hydroxy propyl methyl cellulose (HPMC) and carboxy methyl cellulose (CMC).
A method according to Claim 3; characterized in that the carrier polymer is base chitosan.
A method according to Claim 3; characterized in that the carrier polymer is water soluble chitosan.
A method according to Claim 1; characterized in that the polymers used in creating solid dispersions is selected among cellulose derivatives such as hydroxypropyl methyl cellulose (HPMC), ethyl cellulose, hydroxypropyl cellulose (HPC), sodium carboxymethyl cellulose (NaCMC), starch derivatives, polyvinyl pyrrolidone, eudragit, polyethylene glycols, polymethacrylates, chitosans with various deacetylation, various molecular weights and various solubility, inulin, compritol 888 ATO, gelucire, pluronics, gelatin, cyclodextrins, Kollidon, Avicel 200, Synperonic and Plasdone.
7. A method according to Claim 6; characterized in that the polymer used in creating solid dispersions is polyethylene glycol, pluronic F-68 or base chitosan.
8. A method according to Claim 1; characterized in that the active agent is used at a rate of 0.5-4% (weight/volume).
9. A method according to Claim 1; characterized in that the active agent is used at a rate of 2% (weight/volume).
10. A method according to Claim 1; characterized in that the carrier polymer is used at a rate of 0.1-10% (weight/volume).
1 1 .A method according to Claim 10; characterized in that the carrier polymer is used at a rate of 2% (weight/volume).
12. A method according to Claim 1; characterized in that the active agent:polymer ratio is in the range of 1:3 - 1: 12.
13. A method according to Claim 12; characterized in that the active agent:polymer ratio is 1 :9.
14. A method according to Claim 12; characterized in that the active agent:polymer ratio is 1 : 12.
15.A method according to Claim 1; characterized in that it comprises the process step of preparing the active agent-polymer solid dispersion with the melting method, the co- evaporation method, co-milling method or the kneading method.
16.A method according to Claim 15; characterized in that it comprises the process step of preparing the polymer-active agent solid dispersion with the melting method.
17. A method according to Claim 16; characterized in that the polymer is Pluronic F-68 or PEG 6000 and the active agent is atorvastatin.
18. A method according to Claim 17; characterized in that it comprises the steps of:
- heating the polymer in the water bath at 40-100°C until it melts,
- then, adding the active agent to this melt and constantly mixing until a homogeneous dispersion is obtained,
- storing the mixture inside the refrigerator or the desiccator until solid, and
- then, pulverizing the mixture inside the porcelain mortar.
19.A method according to Claim 18; characterized in that the polymer is heated in the water bath at 58°C until it melts.
20. A method according to Claim 15; characterized in that it comprises the process step of preparing the polymer-active agent solid dispersion with the co-evaporation method.
21 . A method according to Claim 20; characterized in that the polymer is Pluronic F-68 or PEG 6000 and the active agent is atorvastatin.
22.A method according to Claim 21; characterized in that it comprises the steps of:
- dissolving atorvastatin and the polymer in an organic solvent,
- then, evaporating the solvent,
- drying the obtained polymer-atorvastatin mixture until no moisture remains,
- preferably, keeping the mixture inside the desiccator or the refrigerator for 24 hours, - then, pulverizing the mixture inside the porcelain mortar.
23. A method according to Claim 22; characterized in that the organic solvent is methanol.
24. A method according to Claim 23; characterized in that methanol is evaporated in the rotating evaporator at 30-80°C.
25. A method according to Claim 24; characterized in that methanol is evaporated in the rotating evaporator at 40°C.
26. A method according to Claim 15; characterized in that it comprises the process step of preparing the polymer-active agent solid dispersion with the co-milling method.
27. A method according to Claim 26; characterized in that the polymer is base chitosan and the active agent is atorvastatin.
28. A method according to Claim 27; characterized in that the atorvastatin-base chitosan physical mixture is pulverized by mixing for 1-24 hours at 100-600 rpm.
29. A method according to Claim 28; characterized in that the atorvastatin-base chitosan physical mixture is pulverized by mixing for 1 hour at 300 rpm inside the ball mill.
30. A method according to Claim 15; characterized in that it comprises the process step of preparing the polymer-active agent solid dispersion with the kneading method.
31 .A method according to Claim 30; characterized in that the polymer is base chitosan and the active agent is atorvastatin.
32.A method according to Claim 31; characterized in that it comprises the steps of:
- placing the physical mixture of atorvastatin-base chitosan inside the porcelain mortar and mixing said mixture with organic solvent or organic solvent: water mixture, the amount of which is 1 to 10 times amount the weighed powder,
- drying the obtained mixture by keeping it at 10-80°C for 1-5 days.
33. A method according to Claim 32; characterized in that the obtained mixture is dried by keeping it at 50°C for 2 days.
34. A method according to Claim 32; characterized in that the organic solvent: water ratio is in the range of 0.1: 1 - 20: 1.
35. A method according to Claim 34; characterized in that the organic solvent: water ratio is 6: 1.
36. A method according to Claim 35; characterized in that the organic solvent is methanol.
37. A method according to Claim 1; characterized in that the gel composition with base chitosan is prepared inside a diluted acid solution.
38. A method according to Claim 37; characterized in that the diluted acid solution has a concentration of 0.1-10%.
39. A method according to Claim 38; characterized in that the diluted acid solution has a concentration of 1%.
40. A method according to Claim 39; characterized in that the acid inside the diluted acid solution is selected among organic acids such as acetic acid, lactic acid, citric acid, malic acid and formic acid.
41 .A method according to Claim 40; characterized in that the diluted acid solution is a lactic acid solution.
42. A pharmaceutical composition obtained with the method according to any of the above- mentioned claims.
43. The use of a pharmaceutical composition according to Claim 42 in the preparation of a drug for treating periodontal diseases.
44. The use of a pharmaceutical composition according to Claim 42 in the preparation of a drug for treating dermal or mucosal inflammations for topical purposes.
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Citations (3)

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