CREAM CONTAINING CHERENOL-LOADED PLGA NANOPARTICLES FOR TOPICAL USE IN PSORIASIS TREATMENT AND ITS MANUFACTURING METHOD.

Abstract

The invention relates to a nanoformulation and fabrication method involving chirenol-loaded polylactic-co-glycolic acid (PLGA) nanoparticles that provide high skin penetration and a low side effect profile in psoriasis, targeting the S1PR1 signaling axis with ADAM10 and ADAM17 metalloproteases.

Description

1 TARIFF CHERENOL-LOADED PLGA FOR TOPICAL USE IN THE TREATMENT OF PSORIASIS. CREAM CONTAINING NANOPARTICULAR AND ITS PRODUCTION METHOD Technical Area 5 The invention relates to a kirenol-loaded poly(lactic-co-glycolic acid) (PLGA) for the treatment of psoriasis. This invention relates to a topical cream containing a nanoparticle based on a nanoparticle and its production method. The technical aspects of the invention... The field includes nanotechnology-based topical drug development in dermatology. controlled release systems, biodegradable polymer-based drug delivery systems, and Topical pharmaceutical formulations for inflammatory skin diseases 10 It includes. State of the Art Psoriasis is a disease in which the immune system targets the skin and sometimes the joints. Chronic and 15-year-old disease that develops as a result of a combination of genetic predisposition and environmental triggers. It is an inflammatory disease that progresses in attacks (with periods of exacerbation and remission). The pathogenesis of the disease particularly involves the IL-23 / Th17 / IL-17 axis (interleukin-23 / T helper keratinocytes via 17 cells / interleukin-17) and TNF-α (tumor necrosis factor alpha). proliferation accelerates, epidermal thickening and marked inflammation. It consists of 20 sharply demarcated, erythematous (red) plaques. The most typical symptoms are sharply demarcated, erythematous (red) plaques. Silvery-white scales (dandruff), itching / burning, cracking and bleeding, on the scalp. Crusting, pitting of the nail, onycholysis, and in some cases joint pain. It manifests as swelling (psoriatic arthritis). Clinical severity varies from person to person. Psoriasis Although the incidence of the disease varies geographically, it is generally observed in the general population. It is in the 1–3% range. Studies have found the prevalence to be close / similar between women and men, 25 There is evidence that the severity of the disease may be higher in men. Psoriasis The prognosis of the disease is long-term control in most patients. It is being evaluated. With appropriate treatment and trigger management, the lesions are significantly reduced. Although it can regress, the disease tends to recur, so treatments are usually focused on preventing attacks. After being intensified and controlled during the initial period, maintenance or intermittent 30 This is how it is continued. In other words, it has to be applied at the same dose continuously. It is not. However, long-term planning and monitoring are frequently necessary, especially in moderate to severe cases. It is necessary. 2 The treatment of psoriasis depends on the severity of the disease and the affected area (face, genital, scalp, The nails are categorized according to accompanying psoriatic arthritis and comorbidities. In mild cases, topical (local) treatments are the priority. Topical corticosteroids reduce redness. It rapidly reduces itching and inflammation. Vitamin D analogs (e.g., calcipotriol) It is effective in reducing hyperproliferation and plaque thickness. Topical retinoids 5 (Tazarotene) regulates keratinization and is a calcineurin inhibitor. (tacrolimus / pimecrolimus) inflammation, especially in the facial-intertriginous region. It suppresses [the condition]. In moderate / resistant cases, phototherapy reduces plaques and itching. In patients with moderate to severe disease or joint involvement, systemic options are preferred. These systemic options include methotrexate, cyclosporine, and acitretin. This is exemplified by PDE4 inhibitors among small molecules. Apremilast (a phosphodiesterase-4 inhibitor) reduces inflammatory mediators and More targeted immune signal suppressors reduce cytokine signaling. Biological agents are the current group of highly effective treatments for moderate to severe plaque psoriasis. This is due to the fact that TNF-α inhibitors (adalimumab, infliximab, etanercept, etc.) inhibit TNF-15. It suppresses mediated inflammation. However, infection and latent tuberculosis are also present in this group. There is a risk of reactivation. The IL-12 / 23 p40 inhibitor ustekinumab is effective. These are among the options. IL-17 pathway inhibitors can provide rapid plaque clearance. However, there is a risk of mucocutaneous candidiasis infection and in some patients... Use with caution due to the possibility of exacerbation of inflammatory bowel disease. 20 IL-23 p19 inhibitors (guselkumab, risankizumab, tildrakizumab, etc.) are required. By suppressing the IL-23 driver, it indirectly reduces the Th17 response and extends the lifespan. It can provide long-term disease control. Generally, injections in biological classes Reactions and infection risk are key monitoring topics. ADAM10–ADAM17–S1PR1 The current treatment gap is noteworthy in terms of pathways. Of the 25 currently approved treatments... most cytokine axes (TNF-α, IL-23, IL-17, IL-12 / 23) or intracellular signaling (PDE4, (TYK2 etc.) targets. In contrast, although its role in psoriasis has been demonstrated. despite clinically established, safe approaches to metalloproteases such as ADAM17 and ADAM10 and selective drug options are limited. The main reason for this is that ADAM17 targets keratinocytes. its differentiation and its critical functions in skin barrier homeostasis. Broad 30 narrow spectrum inhibition, barrier disruption and undesirable immune effects This can lead to therapeutic window problems. In other words, the target is biologically... Although attractive, striking a balance between dose, selectivity, and safety is difficult. In the S1PR1 axis, however... The mechanism is theoretically meaningful. The retention of lymphocytes in lymphoid tissues and the cells 3 Reducing blood flow can lower inflammation in psoriasis. In this class Ponesimod is an example of an agent studied in psoriasis. However, S1P receptor modulators; bradycardia and AV block, hypertension, macular edema and Caution is required due to systemic safety concerns such as the risk of infection. There is also a risk of relapse if treatment is discontinued. For these reasons, the S1PR1 target is 5. It has not yet become a widely accepted standard axis of care in psoriasis treatment and It remains a relative gap in clinical practice. Patent application CN101597212A, which is included in the prior art, concerns kirenol. synthesis of derivatives and the use of these derivatives, especially proinflammatory agents such as TNF-α and IL-1β 10 a study on its use in the preparation of pharmaceutical agents that inhibit cytokines The document in question describes an invention of a chemical compound and its use in medicine. By focusing on the molecular mechanisms of inflammation-related diseases, TNF-α and It is aimed at identifying new chemical derivatives that target critical cytokines such as IL-1β. And this approach is used for rheumatoid arthritis, autoimmune diseases, and inflammatory diseases. It offers potential therapeutic applications in its treatment. It also includes different substituents. Identification of a broad family of chemical derivatives in terms of structure-activity relationship expanding the research area and alternative candidate molecules for pharmaceutical development. It presents [this]. However, the document's shortcomings include bioavailability and its effect on target tissue. Pharmacokinetic and pharmaceutical performance, such as reach and efficacy in clinical practice 20 The parameters are not discussed in detail. In addition, the current narrative... Targeting ADAM10, ADAM17 and S1PR1 (sphingosine-1-phosphate receptor-1) pathways. its failure to address approaches, particularly alternative mechanisms outside the cytokine axis in terms of expanding assessment and non-responsive / resistant patient strategies This constitutes a significant deficiency. 25 The study by Sun et al., which is part of the state of the art, involved curcumin-loaded... PLGA (poly(lactic-co-glycolic acid)) nanoparticles in a hydrogel formulation Prepared and applied topically in the treatment of psoriasis, it provides skin penetration and therapeutic effects. It is related to an experimental pharmaceutical carrier system that investigates how to increase its effectiveness [1]. 30 The study aimed to solve the problems of curcumin's low solubility and limited stability. To achieve this, PLGA nanoparticles with sizes of 50 nm and 150 nm were developed to deliver the drug to the skin. systematic methods to increase penetration, controlled release and biological efficacy It presents a formulation approach. It also works on a psoriasis-like mouse model. 4 biomarker analysis, histological evaluation and penetration studies were performed. experimentally investigates the topical treatment performance of a nanoparticle delivery system. This indicates that, however, the study has shortcomings regarding the treatment of psoriasis. commonly used topical corticosteroids, calcineurin inhibitors, or biological agents a direct comparative analysis in terms of long-term clinical efficacy and safety 5 It does not offer. Furthermore, it does not offer treatment targeting the ADAM10, ADAM17, and S1PR1 pathways. its absence of concepts regarding targets outside the classic TNF-α–IL-23–IL-17 line It leaves out innovative options. In the current technique, topical corticosteroids and 10, which are commonly used in the treatment of psoriasis, are employed. Long-term administration of calcineurin inhibitors and systemic / biological agents may lead to skin atrophy. clinically significant risks include immunosuppression, increased risk of infection, and systemic toxicity. leading to significant side effects, in contrast to the low bioavailability of natural compounds, Limited efficacy due to photochemical instability and inability to penetrate the skin barrier. Due to shortcomings such as those shown, improvement is needed in this area. 15 It has been made. Brief Description and Objectives of the Invention The invention describes the nano-sized concentration of chyrenol within a poly(lactic-co-glycolic acid) (PLGA) matrix. a cream formulated for topical application and the steps involved in its production 20 It is explained that the cream subject to the invention is ADAM10, ADAM17 and S1PR1. It is designed to produce a therapeutic effect through its axis and in psoriasis It aims to suppress inflammatory processes. The main aim of the invention is to develop a low-side-effect treatment for psoriasis management, skin 25 A nanoformulation based on a natural compound with high penetration and enhanced efficacy. The invention describes the provision of a cream containing chyrenol, a biodegradable diterpene. by nanoencapsulating it within a PLGA matrix, which is a biocompatible polymer. The stability of kirenol is increased, its controlled release is ensured, and it penetrates the skin barrier. The transition is facilitated. Thus, thanks to this invention, the target for psoriasis treatment is 30. Higher local bioavailability in tissue and a safer topical application profile. is obtained. Another aim of the invention is to provide symptom suppression in the treatment of psoriasis. not limited to this approach, but a more targeted approach to the biological processes at the root of the disease. a cream (nanoformulation) whose therapeutic efficacy has been enhanced by providing intervention The aim of this invention is to provide chirenol within a PLGA nano-carrier system. With transport, the formulation contains epidermal proliferation inhibitors such as ADAM10 and ADAM17. and metalloproteases associated with inflammation and their role in regulating the immune response. targeting the S1PR1 (sphingosine-1-phosphate receptor-1) axis This is made possible by its positioning. Thus, the basic pathogenesis of psoriasis More targeted anti-inflammatory effect through modulation of signaling pathways is presented. 10 Another purpose of the invention is to develop it for clinical and industrial use in the treatment of psoriasis. It is highly applicable, characterizable, standardizable, and commercializable. The aim is to provide a nanoformulation (cream) of a certain quality. Within the scope of the invention, kirenol, The nanostructure is loaded into a PLGA matrix and the resulting nanostructure has a size distribution and surface area of ​​15. critical factors such as drug load, morphology, encapsulation efficiency, drug loading rate, release profile, and stability. It can be characterized in terms of quality parameters. This approach, It supports the repeatability and quality assurance of nanoformulation. Thus, in the health, biotechnology, dermatology and dermocosmetics sectors a new topical pharmaceutical product with viable, safe and controlled release characteristics 20 Its development is targeted. Another aim of the invention is to address the issues of chirenol's low water solubility and limited bioavailability. By reducing pharmaceutical disadvantages, the active substance is transported effectively to the target tissue. The aim is to develop a nano-carrier system that provides this. In this invention, a PLGA matrix 25 is used. The stability of kirenol is increased through nanoencapsulation performed within it. its degradation is reduced and its topical pharmaceutical applicability is improved. is being strengthened. Another technical result aimed at by the invention is the controlled and time-dependent 30% absorption of chyrenol. The goal is to ensure its release. This increases the duration of the therapeutic effect. The frequency of application is reduced and patient compliance is supported. In addition The developed nanoformulation allows for more effective penetration of the skin barrier and activates the active targeted accumulation of the compound in the epidermal and dermal layers 6 The aim is to achieve this. In this context, the nano size range, surface properties, and carrier system are being considered. By optimizing its structure, the local therapeutic effect is enhanced. Another aim of the invention is to provide a naturally derived compound as an alternative to synthetic agents. a treatment containing, biocompatible, low toxicity profile and suitable for long-term use 5 The aim is to develop this approach. In this context, the natural bioactive properties of kirenol are being considered. It is used in the treatment of psoriasis and has high patient compliance and can be applied topically. It is offered as an option. Another advantage offered by the invention is that 10 enter the systemic circulation via topical application. The goal is to reduce the amount of active substance. This approach allows for the reduction of existing systemic problems. A safer treatment alternative is offered compared to existing treatments. The goal of this invention is... Another technical effect is that the developed nanoformulation targets CD3⁺ T lymphocytes and MPO⁺ neutrophils. Contributes to the restoration of epidermal immune balance by reducing infiltration. Its purpose is to provide and suppress the accumulation of inflammatory cells in skin lesions. 15 The invention also involves the suppression of Ki-67 positivity and NF-κB p65 phosphorylation. It aims to reduce keratinocyte hyperproliferation specific to psoriasis. This This results in epidermal thickening, parakeratosis, and histopathological features specific to psoriasis. Changes are being reduced. 20 From a systemic safety perspective, the invention protects the liver and kidneys following topical application. to establish a secure profile for the preservation of its functions The aim is to monitor serum ALT, AST, BUN, and creatinine levels. Systemic safety assessment can be performed and the nanoformulation can be tested in the liver for 25 days. Safe use is achieved in terms of kidney function. One of the aims of the invention is the physicochemical development of the nanoformulation. The aim is to increase its stability. This ensures a long shelf life and allows for better storage conditions. obtaining a product that exhibits durable, homogeneous and repeatable quality characteristics. 30 Furthermore, the developed nanoformulation is being transferred from the laboratory scale to the industrial scale. It has a production process that can be scaled up, repeated, and is cost-effective. The aim is to ensure the commercialization of the product and pharmaceutical development. Its transferability to processes is supported. 7 Another platform-related aspect of the invention is the developed PLGA-based nano-carrier. The system offers a structure that can be combined with different active ingredients in the future. This In this respect, the invention is not dependent on a single active substance, but is modifiable and It provides an expandable nano-carrier technology infrastructure. The invention is essentially based on... Although developed for the treatment of psoriasis, it is also used for inflammatory skin conditions. It can be used in the development of topical nano-delivery systems for diseases. It offers a platform technology. Finally, the invention uses the active ingredient chyrenol, which grows naturally in the flora of Türkiye. Because it can be obtained from plants such as Siegesbeckia orientalis, in the long term 10 bioactive that enables the establishment of a local and sustainable production infrastructure It aims to develop a resource-based treatment strategy. Explanation of the Figures Figure 1: 15 of the kirenol-loaded PLGA nanoformulation and topical cream formulation Flow chart showing the production process [A: Organic phase preparation (PLGA+crenol+solvent), B: PVA aqueous phase preparation (1% w / v PVA, 25-80 °C), C: O / W emulsion formation (Sonication, 0.5 min, 30% amplitude), D: Solvent evaporation (25 °C, 4-5 hours, magnetic) E: Centrifugation and washing (20.00g, 20 min, 2 times), F: Vacuum pre-drying (40 C), G: Lyophilization (5% w / v trehalose) and H: Topical cream preparation (20% of the cream base) dispersion, sterilization) Figure 2: Schematic cross-sectional model of chirenol-loaded PLGA nanoparticles (A: PVA stabilizer) A: PLGA polymer matrix, C: Chirenol molecules, and D: Nanoparticle size. range) Detailed Description of the Invention The invention relates to the ADAM10, ADAM17, and sphingosine-1-phosphate receptor-1 (S1PR1) pathways. Developed for topical use in the treatment of psoriasis, targeting therapies that are loaded with kirenol. poly(lactic-co-glycolic acid) (PLGA) nanoformulation (cream) and this It relates to the production method of the nanoformulation (cream). The nanoformulation that is the subject of the invention is 30 Topical application of kirenol: solubility, stability and bioavailability issues It is designed to improve conditions and increase skin penetration, making it more effective. 8 the substance reaches a higher local concentration in the lesion area It provides. The invention targets the ADAM10, ADAM17, and S1PR1 pathways in the treatment of psoriasis (psoriasis). a kirenol-loaded PLGA nanoparticles for use in the treatment of (disease) It is a nanoformulation. Within the scope of the invention, the term "nanoformulation" refers to a 5-cyrenol loaded nanoformulation. PLGA nanoparticles and a carrier suitable for topical application of these nanoparticles. the final pharmaceutical form (cream) obtained by distribution within the system The phrase "kyrenol-loaded PLGA nanoparticles" refers to kyrenol in PLGA polymers. It describes the nano-sized interstitial carrier structure in which the matrix is ​​encapsulated. The carrier system mentioned is the cream base. 10 The final topical cream base in question is hydrophilic, containing chyrenol-loaded PLGA nanoparticles. The oil / water (O / W) type contains a cream base. In one application of the invention, Nanoparticles will be present in the formulation at a concentration of 1% or 2.5% (w / w) chirenol equivalent. It is located in this way. In one application of the invention, hydrophilic water in oil is used. Cream base used as a carrier system with (O / W) emulsion structure (carrier 15 (system), Olus oil, cetearyl alcohol, glyceryl stearate, lecithin, tocopheryl acetate and pharmaceutical acceptable O / W emulsifiers, propylene glycol and paraben-free. O / W is a cream base. In one application of the invention, the aforementioned hydrophilic substance is water in oil (O / W). Cream base (carrier system) used as a carrier system with emulsion structure, 20 - This, - Olus oil as a vegetable-based oil, - Cetearyl alcohol is a fatty alcohol. - PEG-150 Stearate, PEG-75 Stearate, Polysorbate 60, Steareth-20 and Glyceryl O / W emulsifier system containing stearate, 25 - Emollient components include Dicaprylyl Carbonate, Isohexadecane, Isododecane, C13-15 Alkane and humectant (Propanediol), - lecithin as an auxiliary emulsifier, - Polyacrylate-13 and Polyisobutene as stabilizers, - Tocopheryl Acetate as an antioxidant, 30 - Benzoic acid and sorbic acid as a protective system. 9 - Disodium EDTA as a chelating agent) and - It contains sodium hydroxide as a pH regulator. The chyrenol used in this invention is obtained from Siegesbeckia species and It is a naturally occurring diterpenoid known to exhibit anti-inflammatory and antioxidant properties. 5 It acts as the active pharmaceutical ingredient in the formulation. PLGA, on the other hand... It is a biodegradable and biocompatible copolymer that encapsulates and controls the release of chyrenol. It acts as a nano-carrier matrix that enables its release. The polyvinyl alcohol (PVA) used in the formulation described in the invention is used in nanoparticle production. 10 It acts as a surfactant in the process and increases emulsion stability. It promotes the formation of homogeneous nanoparticles. PVA is greatly reduced by washing processes. is largely removed and adsorbed on the nanoparticle surface in the final formulation. It can be found in trace amounts. Trehalose, on the other hand, is detected during the lyophilization process. It is used as a cryoprotectant and after the drying process, nanoparticles 15 It contributes to the preservation of its structure. Therefore, trehalose, when used... Depending on the formulation variant, it is included as an auxiliary component in the final product. is able to obtain. In one application of the invention, the cream, which is a nanoformulation, contains a hydrophilic carrier phase. It is designed as an O / W type emulsion system. This cream base is a topical 20 suitable for application, compatible with skin pH, highly physically stable, and containing nanoparticles. It is a carrier phase that allows for homogeneous distribution. In one application of the invention, this cream some, Fagron Versatile™ Rich O / W Cream Base or equivalent hydrophilic O / W It can be selected as a cream base. The manufacturing method for preparing the nanoformulation described in this invention involves kirenol and 25 The only emulsion-solvent that enables the production of PLGA-containing nanoparticles. It is based on the evaporation technique. The production method of the nanoformulation that is the subject of the invention; i. PLGA and kirenol in dimethyl sulfoxide (DMSO), acetone or ethyl acetate completely dissolved in it with the help of a magnetic stirrer preparation of the organic phase, ii. Distilled containing polyvinyl alcohol (PVA) for use as an aqueous phase. water preparation, 5 iii. Adding the prepared organic phase drop by drop to the aqueous phase and the mixture over an ice bath using a probe sonar by subjecting to sonication, containing kirenol and PLGA an oil / water (O / W) type emulsion consisting of droplets creation, 10 iv. Mixing the resulting emulsion on a magnetic stirrer. and in this process, the organic solvent is removed and PLGA is formed. separation from the solvent and formation of solid nanoparticles obtaining nanoparticle suspension by providing, v. centrifuging the resulting nanoparticle suspension 15 Precipitation of nanoparticles, vi. Removal of supernatant and precipitation of nanoparticles with distilled water. by washing to reduce PVA, free chyrenol and solvent residues, vii. Preliminary drying of the washed nanoparticle suspension under vacuum dryer subjected to drying, 20 viii. Trehalose in nanoparticle suspension subjected to pre-drying lyophilization process in a medium containing and dry powder Obtaining chirenol-loaded PLGA nanoparticles in this form, ix. sterile distillation of the obtained lyophilized chirenol-loaded PLGA nanoparticles suspended in water, 25 through a 0.45 µm membrane filter. by passing it through bioburden reduction filtration and ultrasonic by homogenization process in the bath for 5 minutes ensuring a uniform and homogeneous distribution, x. Hydrophilic, oil / water (O / W) type, cream base suitable for temperatures up to 40–45 °C. heating, 30 xi. the obtained nanoparticle suspension to the heated cream base 1% or corresponding to the targeted active substance dose Slow addition at a rate of 2.5% (w / w) kirenol equivalent, 11 xii. Added suspension is mixed with a magnetic stirrer at 300-500 rpm or For 15–20 minutes with a homogenizer at 1000–3000 rpm. by mixing and ensuring a homogeneous distribution of the kirenol-loaded Hydrophilic topical cream containing PLGA nanoparticles Obtaining (the nanoformulation) 5 It includes the steps involved in the process. In one application of the invention, the method for producing the nanoformulation in question is as follows: i. 25–75 mg PLGA and 2.5–7.5 mg chirenol in 0.5–2 mL DMSO, acetone or in ethyl acetate at 300–700 rpm for 5–20 minutes organic 10 by completely dissolving it with the help of a magnetic stirrer preparation of the phase, ii. 3–7 mL containing 0.5–2% (w / v) PVA for use as the aqueous phase. Preparation of distilled water, iii. The entire prepared organic phase is added dropwise to the aqueous phase. and the temperature that may be generated during sonication of the mixture is 15 kept within a temperature range of 0–10 °C to control its increase 0.25–1 minute over an ice bath using a probe sonicator. by sonicating for a period of time and at an amplitude of 20–40%, kirenol and An oil / water (O / W) type of droplet containing PLGA. Emulsion formation, 20 iv. The resulting emulsion is kept at a temperature range of 20–30 °C for 3–6 hours. stirring on a magnetic stirrer throughout and during this process Separation of PLGA from the solvent by removing the organic solvent. and by enabling the formation of solid nanoparticles, nanoparticle Obtaining the suspension, 25 v. the resulting nanoparticle suspension at 15,000–25,000×g 15–25 Precipitation of nanoparticles by centrifugation for a period of minutes, vi. Removal of supernatant and precipitation of nanoparticles with distilled water. Washing 2–3 times removes PVA, free kirenol and solvent residues. reduction, 30 vii. Preliminary drying of the washed nanoparticle suspension under vacuum dryer subjected to drying, 12 viii. 3–7% (w / v) of nanoparticle suspension subjected to pre-drying Lyophilization process in trehalose-containing medium and drying Preparation of chirenol-loaded PLGA nanoparticles in powder form, ix. sterile distillation of the obtained lyophilized chirenol-loaded PLGA nanoparticles suspended in water, 5 through a 0.45 µm membrane filter by passing it through bioburden reduction filtration and ultrasonic by homogenization process in the bath for 5 minutes ensuring a uniform and homogeneous distribution, x. Hydrophilic, oil / water (O / W) type, cream base suitable for temperatures up to 40–45 °C. heating, 10 xi. the obtained nanoparticle suspension to the heated cream base 1% or corresponding to the targeted active substance dose Slow addition at a rate of 2.5% (w / w) kirenol equivalent, xii. Added suspension is mixed with a magnetic stirrer at 300-500 rpm or 15-20 minutes with a homogenizer at 1000–3000 rpm by mixing and ensuring a homogeneous distribution of the kirenol-loaded Hydrophilic topical cream containing PLGA nanoparticles (obtaining the nanoformulation) It includes the steps involved in the process. In another application of the invention, the production method of the nanoformulation in question is; 20 i. 50 mg PLGA and 5 mg kirenol in 1 mL DMSO, acetone or ethyl acetate magnetic stirring inside at 500 rpm for 10–15 minutes. Preparation of the organic phase by completely dissolving it with the help of, ii. 5 mL of distilled water containing 1% (w / v) PVA for use as the aqueous phase. Preparing the water and bringing it to a temperature of 25 °C, 25 iii. The entire prepared organic phase is added dropwise to the aqueous phase. and the temperature that may occur during sonication of the mixture ice kept at a temperature range of 0–5 °C to control its increase Using a probe sonicator, the solution was applied to the bath for 0.5 minutes. 30 containing kirenol and PLGA, subjected to sonication at 30% amplitude. Formation of an O / W type emulsion consisting of droplets, 13 iv. The resulting emulsion is subjected to magnetic treatment at 25 °C for 4–5 hours. mixing on a mixer and in this process the organic solvent by removing it, PLGA is separated from the solvent and the solid by enabling the formation of nanoparticles Obtaining the suspension, 5 v. the resulting nanoparticle suspension was heated at 20,000×g for 20 minutes Precipitation of nanoparticles by centrifugation for a specified period of time, vi. Removal of supernatant and precipitation of nanoparticles with distilled water. After washing twice, PVA, free chyrenol and solvent residues are removed. reduction, 10 vii. Washing the nanoparticle suspension under a vacuum dryer for 40 minutes. Pre-drying at °C, viii. 5% (w / v) of nanoparticle suspension subjected to pre-drying Lyophilization process in trehalose-containing medium and drying Preparation of chirenol-loaded PLGA nanoparticles in powder form, 15 ix. sterile distillation of the obtained lyophilized chirenol-loaded PLGA nanoparticles suspended in water, through a 0.45 µm membrane filter by passing it through bioburden reduction filtration and ultrasonic by homogenization process in the bath for 5 minutes ensuring a uniform and homogeneous distribution, 20 x. Hydrophilic, oil / water (O / W) type, cream base suitable for temperatures up to 40–45 °C. heating, xi. the obtained nanoparticle suspension to the heated cream base 1% or corresponding to the targeted active substance dose Slow addition at a rate of 2.5% (w / w) kirenol equivalent, 25 xii. Added suspension is mixed with a magnetic stirrer at 300-500 rpm or For 15–20 minutes with a homogenizer at 1000–3000 rpm. by mixing and ensuring a homogeneous distribution of the kirenol-loaded Hydrophilic topical cream containing PLGA nanoparticles (obtaining the nanoformulation) 30 It includes the steps involved in the process. 14 In one application of the invention, the PVA solution is prepared at a concentration of 1% (w / v) and PVA is heated to a temperature range of 60–90 °C in order to dissolve it. The prepared PVA solution is then cooled to 25 °C to obtain the aqueous phase. It is used. In an alternative application, 5 mL of distilled water containing 1% (w / v) PVA is used. It can be prepared directly at 25°C. In one application of the invention, the hydrophilic topical cream base Fagron Versatile™ Rich O / W Cream Base is a similar / equivalent hydrophilic O / W cream base. This cream base... to ensure homogeneous distribution of the nanoparticle suspension It is heated to 40–45 °C during the production process. The cream base is ready for topical application in 10 minutes. possessing suitable physical properties, the carrier phase and cream matrix in the final formulation. He / She serves as follows: The final composition of the topical nanoformulation (cream) prepared within the scope of the invention is as follows: - Chirenol-loaded PLGA nanoparticles containing 1% or 2.5% (w / w) chirenol equivalent, - Hydrophilic O / W cream base, qs 100 g, 15 - trace amounts of PVA residue resulting from the production process, - trehalose resulting from the lyophilization process, - sufficient quantity of sterile distilled water used for resuspension It includes its components. The production method described in this invention involves homogeneously dispersing 20 centrifuges in a PLGA matrix. encapsulation, obtaining nano-sized and reproducible particle structures, a cream that ensures controlled release of the active ingredient and is suitable for topical application. This makes it possible to transfer it to the formulation. Organic phase preparation, containing PVA. aqueous phase, sonication, removal of organic solvent, centrifugation-washing, pre-treatment When the drying and lyophilization steps are considered together, the method; particle 25 size, encapsulation efficiency, physical stability, and resuspendability This provides a technical advantage in this respect. The resulting chirenol-loaded PLGA nanoparticles; particle size (target range: 100–250 (nm), zeta potential (target range: –20 to –30 mV), polydispersity index, encapsulation It is characterized in terms of efficacy, drug loading rate, and in vitro release profile. 30 Capsulation yield and drug loading rate are determined by the amount of supernatant remaining after centrifugation. Free chyrenol concentration measured with a UV-Vis spectrophotometer at 210–215 nm wavelength. This is determined by measuring the neck and calculating the result using appropriate formulas. In vitro release profile, dialysis membrane in pH 5.5 acetate buffer at 37 °C. This is done by monitoring the cumulative emission percentage over a period of 0–72 hours. 5 Topical application is achieved by transferring nanoparticles to a hydrophilic O / W cream base. The final nano-cream formulation is obtained; this formulation is the visual phase. in terms of separation, viscosity, content uniformity, short-term stability and release profile. is being evaluated. The biological efficacy of the invention is being evaluated through in vitro and in vivo models. 10 In the in vitro stage, cell viability in the LPS-induced HaCaT cell model was measured using CCK-8. The levels of IL-6, IL-1β, TNF-α, IL-17A, IL-22, and IL-23 were determined using the ELISA method. The expression levels of ADAM10, ADAM17, and S1PR1 proteins were determined using the Western blot method. S1P levels are being examined using the Western blot method due to its lipid structure. It cannot be measured and is determined in cell lysates by ELISA method. 15 In vivo, a psoriasis-like model was developed in C57BL / 6J mice using imiquimod. is being created; histopathological examination of skin tissues, CD3, MPO, Ki-67 and p-NF- κB p65 immunohistochemistry, cytokine levels, ADAM10, ADAM17, SPHK1 and S1PR1 S1P levels are assessed by their expression. Serum ALT, AST, BUN and The systemic safety profile is examined using creatinine measurements. 20 In conclusion, the invention is obtained by encapsulating kirenol within PLGA nanoparticles. Presentation of nanoparticles in topical cream form, ADAM10, ADAM17 in psoriasis and targeting the inflammatory process via the S1PR1 axis and formulation Kirenol, which allows evaluation using physicochemical, in vitro and in vivo methods. It presents a charged PLGA nanoformulation and production method. 25 16 REFERENCES [1] Lin Sun, Zeyu Liu, Lun Wang, Dongmei Cun, Henry HY Tong, Ru Yan, Xin Chen, Ruibing Wang, Ying Zheng, Enhanced topical penetration, system exposure and anti- psoriasis activity of two particle-sized, curcumin-loaded PLGA nanoparticles in hydrogel. 5 15 25

Claims

17 REQUESTS 1. ADAM10 and ADAM17 for use in the treatment of psoriasis. a hydrophilic topical treatment targeting the S1PR1 signaling axis with metalloproteases It is a cream and its characteristic is that it contains chyrenol-loaded poly(lactic-co-glycolic acid) (PLGA) 5. It contains nanoparticles and a carrier system.

2. A hydrophilic topical medication according to Claim 1 for use in the treatment of psoriasis. It is a cream and its characteristic feature is that it contains hydrophilic, oil / water (O / W) particles loaded with chirenol-based PLGA nanoparticles. The type is that the ingredients are evenly distributed within a cream base.

3. A hydrophilic topical 10 according to Claim 2 for use in the treatment of psoriasis. It is a cream and its characteristic is that the cream base in question... - This, - Olus oil as a vegetable-based oil, - Cetearyl alcohol is a fatty alcohol. - PEG-150 Stearate, PEG-75 Stearate, Polysorbate 60, Steareth-20 and Glyceryl 15 O / W emulsifier system containing stearate, - Dicaprylyl Carbonate and Isohexadecane as emollient components, Isododecane, C13-15 Alkane and humectant (Propanediol), - lecithin as an auxiliary emulsifier, - Polyacrylate-13 and Polyisobutene as stabilizers, 20 - Tocopheryl Acetate as an antioxidant, - Benzoic acid and sorbic acid as a protective system. - Disodium EDTA as a chelating agent) and - Sodium hydroxide as a pH regulator. It includes. 25 4. ADAM10 and ADAM17 for use in the treatment of psoriasis. metalloproteases targeting the S1PR1 signaling axis from any of Claims 1-3 According to one source, chirenol-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles It is a production method of a hydrophilic topical cream containing; its characteristic feature is: i. PLGA and kirenol in dimethyl sulfoxide (DMSO), acetone or ethyl acetate 30 by completely dissolving it in it with the help of a magnetic stirrer preparation of the organic phase, 18 ii. Distilled water containing polyvinyl alcohol (PVA) for use as an aqueous phase. preparation, iii. Adding the prepared organic phase drop by drop to the aqueous phase and mixing Sonication is performed over an ice bath using a probe sonicator. oil / water (O / W) 5 formed from droplets containing kirenol and PLGA upon retention. the formation of a type of emulsion, iv. Mixing the resulting emulsion on a magnetic stirrer and this In the process, the organic solvent is removed, separating PLGA from the solvent. and by enabling the formation of solid nanoparticles, nanoparticle Obtaining the suspension, 10 v. by centrifuging the resulting nanoparticle suspension Precipitation of nanoparticles, vi. Removal of supernatant and precipitation of nanoparticles with distilled water. by washing to reduce PVA, free chyrenol and solvent residues, vii. Pre-drying the washed nanoparticle suspension under vacuum dryer 15 subjected to drying, viii. pre-drying of the nanoparticle suspension containing trehalose lyophilization process in the medium and kirenol in dry powder form Obtaining charged PLGA nanoparticles, ix. The obtained lyophilized chirenol-loaded PLGA nanoparticles were mixed with sterile distilled water 20 suspended inside, passed through a 0.45 µm membrane filter subjected to bioburden reduction filtration and 5 minutes in an ultrasonic bath. by subjecting it to a homogenization process for a period of time, resulting in a smooth and homogeneous mixture. ensuring distribution, x. hydrophilic, oil / water (O / W) type, cream base heated to 40–45 °C, 25 xi. The resulting nanoparticle suspension is applied to the heated cream base at the target. Kirenol at 1% or 2.5% (w / w) according to the active ingredient dose. by slowly adding in an equivalent ratio, xii. Added suspension is mixed with a magnetic stirrer at 300-500 rpm or 30 by mixing and ensuring a homogeneous distribution of cirnol-loaded PLGA hydrophilic topical cream (nanoformulation) containing nanoparticles is obtained to be done 19 It includes the steps of the process.

5. ADAM10 and ADAM17 for use in the treatment of psoriasis. nanoformulation targeting the S1PR1 signaling axis with metalloproteases The production method of the chyrenol-loaded PLGA nanoparticles it contains is in accordance with Claim 4. Feature; 5 i. 25–75 mg PLGA and 2.5–7.5 mg kirenol mixed with 0.5–2 mL DMSO, acetone, or Magnetic in ethyl acetate at 300–700 rpm for 5–20 minutes. the organic phase is completely dissolved with the help of a mixer preparation, ii. 3–7 mL of distilled water containing 0.5–2% (w / v) PVA for use as the aqueous phase. water preparation, iii. Adding the entire prepared organic phase to the aqueous phase drop by drop. and control the temperature increase that may occur in the mixture during sonication. on an ice bath kept at a temperature range of 0–10 °C for the purpose of... A probe sonicator is used for 0.25–1 minute at an amplitude of 20–40% for 15 minutes. from droplets containing kirenol and PLGA by subjecting them to sonication the formation of an oil / water (O / W) type emulsion, iv. The resulting emulsion is kept at a temperature range of 20–30 °C for 3–6 hours. mixing on a magnetic stirrer and organic in this process Separating PLGA from the solvent by removing the solvent and solid 20 by enabling the formation of nanoparticles, nanoparticle suspension obtaining, v. the resulting nanoparticle suspension is heated at 15,000–25,000×g for 15–25 minutes. Precipitation of nanoparticles by centrifugation for a specified period of time, vi. Removal of supernatant and rinsing of precipitated nanoparticles with distilled water 2–3 25 by washing several times to reduce PVA, free kirenol and solvent residues, vii. Preliminary drying of the washed nanoparticle suspension under vacuum dryer subjected to drying, viii. 3–7% (w / v) of nanoparticle suspension subjected to pre-drying Lyophilization process in trehalose-containing medium and dry powder 30 Obtaining chirenol-loaded PLGA nanoparticles in this form, ix. The resulting lyophilized chirenol-loaded PLGA nanoparticles were mixed with sterile distilled water. suspended inside, passed through a 0.45 µm membrane filter subjected to bioburden reduction filtration and 5 minutes in an ultrasonic bath. by subjecting it to a homogenization process for a period of time, resulting in a smooth and homogeneous mixture. ensuring distribution, x. Heating of hydrophilic, oil / water (O / W) type cream base to 40–45 °C, xi. The resulting nanoparticle suspension is applied to the heated cream base at the targeted 5 Kirenol at 1% or 2.5% (w / w) according to the active ingredient dose. by slowly adding in an equivalent ratio, xii. Added suspension is mixed with a magnetic stirrer at 300-500 rpm or For 15–20 minutes with a homogenizer at 1000–3000 rpm. by mixing and ensuring a homogeneous distribution of plegas 10 loaded with chyrenol. hydrophilic topical cream (nanoformulation) containing nanoparticles is obtained to be done It includes the steps of the process.

6. ADAM10 and ADAM17 for use in the treatment of psoriasis. 15 nanoformulations targeting the S1PR1 signaling axis with metalloproteases The production method of the chyrenol-loaded PLGA nanoparticles it contains is in accordance with Claim 5. feature; i. 50 mg PLGA and 5 mg kirenol in 1 mL DMSO, acetone or ethyl acetate magnetic stirring inside at 500 rpm for 10–15 minutes. Preparation of the organic phase by completely dissolving it with the help of, 20 ii. 5 mL of distilled water containing 1% (w / v) PVA to be used as the aqueous phase. preparation and bringing the temperature to 25 °C, iii. Adding the entire prepared organic phase to the aqueous phase drop by drop. and control the temperature increase that may occur in the mixture during sonication. For this purpose, 25 on an ice bath kept at a temperature range of 0–5 °C. The probe is sonicated for 0.5 minutes at 30% amplitude using a sonicator. an O / W type of droplet containing kirenol and PLGA, subjected to formation of emulsion, iv. The resulting emulsion is subjected to magnetic treatment at 25 °C for 4–5 hours. mixing on a mixer and in this process 30 organic solvent by removing and separating PLGA from the solvent and solid nanoparticles Obtaining a nanoparticle suspension by ensuring its formation, 21 v. the resulting nanoparticle suspension was heated at 20,000×g for 20 minutes. Precipitation of nanoparticles by centrifugation, vi. Removal of supernatant and rinsing of precipitated nanoparticles with distilled water twice. by washing to reduce PVA, free chyrenol and solvent residues, vii. Washing the nanoparticle suspension under a vacuum dryer at 40 °C for 5 minutes. pre-drying viii. Pre-dried nanoparticle suspension containing 5% (w / v) trehalose lyophilization process in a medium containing and in dry powder form Synthesis of chirenol-loaded PLGA nanoparticles, ix. The obtained lyophilized chirenol-loaded PLGA nanoparticles were mixed with sterile distilled water 10 suspended inside, passed through a 0.45 µm membrane filter subjected to bioburden reduction filtration and 5 minutes in an ultrasonic bath. by subjecting it to a homogenization process for a period of time, resulting in a smooth and homogeneous mixture. ensuring distribution, x. hydrophilic, oil / water (O / W) type, cream base heated to 40–45 °C, 15 xi. The resulting nanoparticle suspension is applied to the heated cream base at the target. Kirenol at 1% or 2.5% (w / w) according to the active ingredient dose. by slowly adding in an equivalent ratio, xii. Added suspension is mixed with a magnetic stirrer at 300-500 rpm or 20 by mixing and ensuring a homogeneous distribution of cirnol-loaded PLGA hydrophilic topical cream (nanoformulation) containing nanoparticles is obtained to be done It includes the steps of the process.

7. Psoriasis 25 obtained by a method according to any of claims 4-6. for use in treatment with ADAM10 and ADAM17 metalloproteases Containing chirenol-loaded PLGA nanoparticles targeting the S1PR1 signaling axis. Hydrophilic topical cream.

8. According to claim 7, it is a hydrophilic topical cream, the characteristic of which is derived from the manufacturing process. trace amounts of PVA residue that may result from the lyophilization process 30 This is due to the fact that it contains trehalose. .

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