Ciclopirox olamine composite medicine and application thereof in preparation of medicine for inhibiting scar

Abstract

The invention provides a ciclopirox olamine composite medicine and application thereof in preparation of a medicine for inhibiting scars, and a preparation method of the ciclopirox olamine composite medicine comprises the following steps: S1, preparing a hyaluronic acid solution; and S2, adding the ciclopirox olamine into a hyaluronic acid solution, and carrying out ultrasonic dissolution to obtain the ciclopirox olamine composite medicine. The ciclopirox olamine composite medicine has the effect of inhibiting hypertrophic scars at lower concentration.

Description

Technical Field

[0001] The invention relates to a ciclopirox compound drug and application thereof in the preparation of a scar inhibiting drug, belonging to the technical field of scar inhibiting drugs. Background Art

[0002] Pathological scars are a type of fibrous metabolic disease characterized by excessive fibrosis and collagen deposition in the human dermis. They are divided into hypertrophic scars and keloids. Scars are abnormal results of skin wound healing, usually caused by burns, trauma or surgery. In severe cases, they can lead to local dysfunction and limited movement in patients, and are often accompanied by related symptoms such as pain, itching, and paresthesia. They not only affect the appearance, but also cause serious psychological and physiological effects, and also bring a huge burden to medical work. With the improvement of people's quality of life, the pursuit of beauty has become stronger. The study of scars has received more and more attention and is a difficult problem that needs to be solved urgently in dermatology and even the entire field of wound repair. The pathogenesis of pathological scars is complex, and genetic factors, extracellular matrix, immune factors, cytokines, and cell apoptosis are all involved. Its treatment strategy mainly uses corticosteroids, scar repair surgery, cryotherapy, radiotherapy, 5-fluorouracil, and some new therapies such as mesenchymal stem cells and interferon. Although there are many treatment methods, there are drug resistance and recurrence. Patients need to repeat treatment or choose surgical resection and adjuvant radiotherapy, which not only brings pain to patients, but also has limited therapeutic effects. Therefore, there is an urgent need for an effective method to improve the treatment effect of scars.

[0003] Ciclopirox belongs to a class of hydroxypyridone antifungal drugs, which has been widely used to treat superficial fungal infections of the skin and nails for three decades, with good efficacy and tolerance. In addition, clinical studies have shown that Ciclopirox has good therapeutic indications, and the safety of Ciclopirox has been demonstrated by evaluating the dose tolerance, pharmacokinetics and pharmacodynamics of patients with refractory malignant hematological diseases. More and more evidence shows that the drug also has anticancer activity against a series of human tumors, including leukemia, lymphoma, myeloma, Ewing's sarcoma, colon adenocarcinoma, cervical cancer, renal cell carcinoma, esophageal cancer, bladder cancer, rhabdomyosarcoma, breast cancer and pancreatic ductal adenocarcinoma, and also has considerable potential to combat many diseases including diabetes, cardiovascular disease and acquired immunodeficiency syndrome. Studies have shown that the anticancer activity of Ciclopirox is related to its inhibition of cell proliferation, induction of apoptosis, inhibition of cell migration and invasion, and inhibition of lymphangiogenesis. At the same time, studies have found that the drug also has the effect of inhibiting angiogenesis and anti-myofibroblasts. However, there is no report on its inhibition of scar hyperplasia. Summary of the invention

[0004] The present invention provides a ciclopirox compound drug and application thereof in the preparation of a scar inhibiting drug, which can effectively solve the above-mentioned problems.

[0005] The present invention is achieved in that:

[0006] A method for preparing a ciclopirox compound drug comprises the following steps:

[0007] S1, preparing hyaluronic acid solution;

[0008] S2, adding ciclopirox to the hyaluronic acid solution and performing ultrasonic dissolution to obtain the ciclopirox composite drug.

[0009] In some embodiments, the concentration of the hyaluronic acid solution is 0-0.1 mg / mL.

[0010] In some embodiments, the concentration of ciclopirox in the ciclopirox combination drug is 2.5-100 μg / mL.

[0011] In some embodiments, the ultrasonic dissolution time is 5-10 min.

[0012] In some embodiments, a pharmaceutically acceptable excipient is also included.

[0013] A ciclopirox compound drug prepared by the above method.

[0014] A use of the above-mentioned ciclopirox compound drug in the preparation of a scar inhibiting drug.

[0015] In some embodiments, the scar is a hypertrophic scar.

[0016] The beneficial effects of the present invention are:

[0017] The ciclopirox compound drug provided by the present invention can simultaneously inhibit the activity and migration ability of human keloid fibroblasts HKF and vascular endothelial cells HUVEC at low concentrations, and has the effect of inhibiting hypertrophic scars at relatively low concentrations.

[0018] The present invention utilizes physical ultrasound to prepare the ciclopirox compound drug, and its preparation process is a purely physical process, during which no other cosolvents, emulsions or other agents that may cause toxicity in the body are introduced. The prepared ciclopirox compound drug has a three-dimensional network structure, and physical ultrasonic mixing successfully forms amide bonds, which can achieve drug retention and has the potential to slow down drug release. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for use in the embodiments will be briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without paying creative work.

[0020] Figure 1 This is a scanning electron micrograph of pure hyaluronic acid in Comparative Example 1.

[0021] Figure 2 This is a scanning electron microscope image of the ciclopirox compound drug of Example 1.

[0022] Figure 3 This is the Fourier infrared spectrum of the ciclopirox compound drug of Example 1.

[0023] Figure 4 The figure is the ultraviolet absorption spectrum of the ciclopirox compound drug of Example 1.

[0024] Figure 5 The figure shows the comparison of cell activity rates of human keloid fibroblasts HKF from Example 2 under different concentrations of ciclopirox.

[0025] Figure 6 The figure is a comparison of the cell activity rates of the vascular endothelial cells HUVEC in Example 2 under different concentrations of ciclopirox.

[0026] Figure 7 The figure is a comparison of the cell migration ability of human keloid fibroblasts HKF in Example 3 under different concentrations of ciclopirox.

[0027] Figure 8 The figure is a comparison of the cell migration ability of the vascular endothelial cells HUVEC in Example 3 under different concentrations of ciclopirox.

[0028] Fig. 9 This is the scar recovery status of the rabbit ear hypertrophic scar model in Example 4 under three different treatments. DETAILED DESCRIPTION

[0029] In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention. Therefore, the following detailed description of the embodiments of the present invention provided in the drawings is not intended to limit the scope of the invention claimed for protection, but merely represents the selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0030] The present invention provides a method for preparing a ciclopirox compound drug, comprising the following steps:

[0031] S1, preparing hyaluronic acid solution;

[0032] S2, adding ciclopirox to the hyaluronic acid solution and performing ultrasonic dissolution to obtain the ciclopirox composite drug.

[0033] First, let's take a look at the preparation process of hyaluronic acid solution. Hyaluronic acid (HA) is a natural high-molecular polysaccharide with good biocompatibility and biodegradability. Hyaluronic acid is an important component of the cell matrix and various tissues. It has important physiological functions such as regulating cell proliferation, migration and differentiation, moisturizing, lubricating, regulating inflammatory response and immunity, and promoting wound healing. In scar treatment, hyaluronic acid can inhibit the formation of scars by inhibiting the differentiation and maturation of fibroblasts and improving collagen arrangement. As a clinical drug for the treatment of scars, it can significantly inhibit collagen proliferation and angiogenesis, while increasing the local retention time of the drug, achieving the effect of reshaping scars in multiple ways. When preparing hyaluronic acid solution, it is necessary to select high-quality raw materials and dissolve them in water by appropriate methods. This process requires strict control of the concentration, pH value, temperature and other conditions of the solution to ensure that the hyaluronic acid can be fully dissolved and maintain its biological activity.

[0034] Next, we will introduce the preparation method of the ciclopirox compound. In this step, ciclopirox is added to a hyaluronic acid solution and ultrasonically dissolved. Ciclopirox (CPX) is a compound with a wide range of pharmacological activities that can be used to treat a variety of diseases, such as inflammation, tumors, etc. By combining with hyaluronic acid, ciclopirox can improve its bioavailability and therapeutic effect in the body. At the same time, the prepared ciclopirox compound successfully formed an amide bond. The formation of this chemical bond means that the drugs interact with each other during the compounding process and have the potential for synergistic enhancement. In addition, ultrasonic dissolution helps ciclopirox dissolve in the solution faster, thereby improving the preparation efficiency of the compound drug.

[0035] In some embodiments, the concentration of the hyaluronic acid solution is 0-0.1 mg / mL.

[0036] In some embodiments, the concentration of ciclopirox in the ciclopirox combination drug is 2.5-100 μg / mL.

[0037] Ciclopirox has a low solubility, and direct clinical application may lead to problems such as low drug bioavailability and unstable efficacy. To solve this problem, the researchers combined Ciclopirox with hyaluronic acid solution to form a new type of compound drug.

[0038] In this compound medicine, the concentration of ciclopirox is crucial. After repeated experimental studies, we find that in certain embodiments, the concentration of ciclopirox in the compound medicine is controlled within the range of 5-400 μM, which can give full play to its pharmacological action, while avoiding the side effect caused by excessive or too low dosage. In addition, the concentration of hyaluronic acid solution also needs to be properly controlled, generally within the range of 0-0.1 mg / mL, to ensure the stability and bioavailability of the drug.

[0039] Through the above research, we have obtained the optimal concentration ratio of ciclopirox and hyaluronic acid solution compound drugs. This discovery provides an important basis for clinical application and helps to improve the efficacy and safety of ciclopirox in the treatment of diseases.

[0040] In some embodiments, the ultrasonic dissolution time is 5-10 min.

[0041] In some embodiments, a pharmaceutically acceptable excipient is also included.

[0042] A ciclopirox compound drug prepared by the above method.

[0043] A use of the above-mentioned ciclopirox compound drug in the preparation of a scar inhibiting drug.

[0044] Example 1: Preparation of Ciclopirox Compound

[0045] (1) Dissolve 4 mg of hyaluronic acid in 40 ml of ultrapure water to obtain a hyaluronic acid solution with a concentration of 0.1 mg / mL.

[0046] (2) Weigh ciclopirox amine and add it to the hyaluronic acid solution obtained in step (1), and dissolve it by physical ultrasonication for 5-10 minutes to obtain a ciclopirox amine composite drug. The concentration of ciclopirox amine in the ciclopirox amine composite drug is 18.5 μM.

[0047] Comparative Example 1

[0048] 4 mg of hyaluronic acid was dissolved in 40 ml of ultrapure water to obtain a hyaluronic acid solution with a concentration of 0.1 mg / mL.

[0049] The results showed the morphological differences between pure hyaluronic acid and ciclopirox in the comparative experiment under scanning electron microscopy. Figure 1 As shown in Figure 2, the pure hyaluronic acid sample showed a smooth surface without obvious pores under a scanning electron microscope. This phenomenon indicates that the pure hyaluronic acid sample is relatively uniform in structure and has no obvious defects.

[0050] Figure 2 The scanning electron microscope image of the ciclopirox amine composite drug in Example 1 is shown. As can be seen from the figure, the composite drug presents a porous three-dimensional network structure. This special structure is conducive to the effective retention of the drug in the body, thereby having the potential to slow down the release of the drug. This means that through this special structural design, the sustained effect of the drug in the body can be achieved, improving the efficacy of the drug.

[0051] Figure 3 and Figure 4 It is respectively the Fourier infrared spectrum and the ultraviolet absorption spectrum of the ciclopirox amide compound drug of embodiment 1.These two drawings show that the prepared ciclopirox amide compound drug successfully forms an amide bond.The formation of this chemical bond means that the drug interacts with each other during the compounding process, and has the potential of synergistic enhancement.In addition, the ultraviolet absorption spectrum also shows that the ciclopirox amide compound drug has certain light absorption performance, and it is expected to realize the release and effect of the drug under illumination conditions.

[0052] Example 2 Comparison of the cell activity rates of drugs on different cells

[0053] Ciclopirox solutions of different concentrations (12.5, 25, 50, 100, and 200 μM, respectively) were prepared.

[0054] Human keloid fibroblasts HKF (Human Keloid Fibroblasts, iCell / Sebikang, from Nanjing Herb Source Biotechnology Co., Ltd.) or human umbilical vein endothelial cells HUVEC (from ATCC) were inoculated into 96-well plates, placed in a cell culture incubator for 12 hours, and then washed twice with DPBS. After the cells were incubated with different concentrations of ciclopirox compound drugs for 12 hours, CCK-8 solution was added to each well and incubated for another 2 hours, the absorbance (OD) value was read at a wavelength of 450nm in an ELISA reader, and the cell activity rate was calculated.

[0055] The experimental results are as follows Figure 5 and Figure 6 As shown, the relationship between the concentration of cyclopyrisamide and the cell proliferation activity is clearly presented. The experimental results show that as the concentration gradient of cyclopyrisamide increases, the proliferation activity of HKF and HUVEC cells gradually decreases. This phenomenon shows that cyclopyrisamide has a concentration-dependent effect on inhibiting the proliferation of HKF and HUVEC cells. In this regard, we conducted a statistical analysis, and the results showed that when the significance level was 0.01 and 0.001, the effect of cyclopyrisamide on inhibiting the proliferation of HKF and HUVEC cells was statistically significant. Cyclopyrisamide has the effect of inhibiting the proliferation of HKF and HUVEC cells, indicating that the cyclopyrisamide compound drug also has the effect of inhibiting the proliferation of HKF and HUVEC cells.

[0056] This discovery is of great significance for a deeper understanding of the biological effects of cyclopyrithamide, and provides a theoretical basis for further exploring the mechanism of action of cyclopyrithamide in different types of cells and disease models in the future, and for clinical application. At the same time, we should also pay attention to studying the biological effects of cyclopyrithamide at low concentrations, in order to provide new ideas for finding new treatment methods.

[0057] Example 3 Comparison of the cell migration ability of different cells by drugs

[0058] Ciclopirox solutions of different concentrations (0, 100 and 200 μM, respectively) were prepared.

[0059] Human keloid fibroblasts HKF or human umbilical vein endothelial cells HUVEC were seeded into 6-well plates, cultured in a cell culture incubator for 12 hours, washed twice with DPBS, and vertically streaked on the bottom wall of each well. The cells were incubated with different concentrations of ciclopirox amide compound drugs, and the migration of human keloid fibroblasts HKF or human umbilical vein endothelial cells HUVEC was observed and photographed under an inverted phase contrast microscope at 0h, 12h, and 24h, and the scratch area was calculated using Image J software.

[0060] The experimental results are as follows Figure 7 and Figure 8As shown, the role of cyclopyrisulfide in cell migration was clearly revealed.

[0061] First, we set up a control group and an experimental group, and the two groups of cells had the same cell migration ability at the beginning of the experiment. Then, we applied cyclopyrisamide to the cells in the experimental group to observe its effect on cell migration ability. The experimental results showed that after 12 hours of drug action, the migration ability of the cells in the experimental group was significantly weakened compared with the control group. This result shows that cyclopyrisamide can inhibit the migration of HKF and HUVEC cells to a certain extent.

[0062] Then, we further extended the drug action time to 24 hours. At this time, the healing speed of the cells in the control group was significantly accelerated, and they almost merged into a piece. However, under the action of cyclopyrisamide, the healing speed of the cells in the experimental group was relatively slow. This result further confirmed the inhibitory effect of cyclopyrisamide on cell migration, and this effect was positively correlated with the drug concentration. Cyclopyrisamide has an inhibitory effect on cell migration, indicating that the cyclopyrisamide compound drug also has an inhibitory effect on cell migration.

[0063] Through statistical analysis, we obtained significant differences in data results. Figure 8 As shown, the difference in cell migration ability between the experimental group and the control group was statistically significant (**p<0.01, ***p<0.001, ****p<0.0001). This indicates that ciclopirox has a significant effect on the migration ability of HKF and HUVEC cells, and this inhibitory effect is more obvious with the increase of drug concentration.

[0064] Example 4 Drugs for the treatment of rabbit ear scars

[0065] (1) According to the method of Example 1, a ciclopirox compound drug having a hyaluronic acid concentration of 10 mg / mL and a ciclopirox concentration of 5.4 mg / mL was prepared.

[0066] (2) A rabbit ear ventral scar model was established by surgical method. After the model was successfully prepared, each scar tissue was injected with the same volume (100 μL) of hyaluronic acid solution (hyaluronic acid concentration was 10 mg / mL) and cyclopyrisamide compound drug aqueous solution. Scars without any treatment served as the control group. The injections were repeated 4 times with an interval of 1 week. The scar efficacy before and after the injection was evaluated one week after the 4th injection.

[0067] The experimental results are as follows Fig. 9As shown, there are significant differences in the treatment of scars among the ciclopirox combination drug treatment group, the scar model group (control group), and the hyaluronic acid solution treatment group. The scar condition of the group treated with the ciclopirox combination drug was significantly better than that of the other two groups, with the scar being flatter, lighter in color, and closer to the surrounding normal skin. This result shows that the ciclopirox combination drug has a significant effect in the treatment of scars.

[0068] In contrast, the scar model group and the hyaluronic acid solution treatment group still showed obvious scar hyperplasia, and the scar color was pink. Although the scar color of the hyaluronic acid solution treatment group was slightly lighter than that of the scar model group, there was still a certain gap compared with the cyclopyrisamide combination drug treatment group. This shows that the hyaluronic acid solution has a certain effect in inhibiting scar hyperplasia, but there is still a certain gap compared with the cyclopyrisamide combination drug.

[0069] In the rabbit ear scar model, the study found that the ciclopirox compound had the effect of inhibiting hypertrophic scars at a lower concentration, overcoming the low solubility of ciclopirox, which may lead to low bioavailability and unstable efficacy when directly used in clinical practice. This shows that ciclopirox and hyaluronic acid play a synergistic role in inhibiting hypertrophic scars. Combined with the above experimental results, it can be considered that the ciclopirox compound has a higher efficacy in treating scars.

[0070] In conclusion, through this experiment, we found that the ciclopirox compound has obvious advantages in the treatment of scars. It not only shows a high effect in inhibiting scar hyperplasia, but also can achieve this effect at a lower concentration. This provides a new possibility for the clinical treatment of scars, which deserves further research and discussion. In future studies, we can further explore the mechanism of action of ciclopirox and hyaluronic acid in inhibiting scar hyperplasia, in order to provide more theoretical basis and practical guidance for scar treatment.

[0071] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for preparing a ciclopirox compound, characterized in that: The following steps are involved: S1, prepare hyaluronic acid solution; S2, adding ciclopirox to the hyaluronic acid solution and performing ultrasonic dissolution to obtain the ciclopirox compound drug.

2. The method for preparing the ciclopirox compound according to claim 1, wherein The concentration of the hyaluronic acid solution is 0-0.1 mg / mL.

3. The method for preparing the ciclopirox compound according to claim 1, wherein In the ciclopirox compound drug, the concentration of ciclopirox is 2.5-100 μg / mL.

4. The method for preparing the ciclopirox compound according to claim 1, wherein The ultrasonic dissolution time is 5-10 minutes.

5. The method for preparing the ciclopirox compound according to claim 1, wherein It also includes the addition of pharmaceutically acceptable excipients.

6. A ciclopirox compound prepared by the method according to any one of claims 1 to 5.

7. Use of the ciclopirox compound according to claim 6 in the preparation of a scar inhibiting drug.

8. The use according to claim 7, characterized in that The scar is a hypertrophic scar.

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