Application of CCL17 as target spot in preparation of medicine for preventing and treating psoriasis

Abstract

The invention discloses application of CCL17 as a target spot in preparation of a medicine for preventing and treating psoriasis, and belongs to the technical field of medicines. The invention provides the application of CCL17 as a target to the medicine for preventing and treating psoriasis for the first time. Experiments show that by inhibiting CCL17, mouse psoriasis induced by imiquimod is remarkably improved, skin lesion symptoms and pathological manifestations of psoriasis mice are relieved, and the expression level of skin lesion inflammatory factors is reduced. The invention proves that the targeted CCL17 can be used as an effective target for relieving psoriasis, provides application potential of the targeted CCL17 in treatment of psoriasis, and has important significance in prevention and treatment of psoriasis.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to the application of CCL17 as a target in the preparation of drugs for preventing and treating psoriasis. Background Art

[0002] Psoriasis, also known as psoriasis, is an immune-mediated chronic, inflammatory, recurrent, systemic skin disease, which was once listed as one of the top ten intractable diseases of mankind by the World Health Organization. Its typical clinical symptoms are red papules or plaques covered with multiple layers of silvery white scales, and its pathological features are hyperkeratosis of the epidermal stratum corneum, thickening of the spinous layer, hyperplasia of the basal layer, and infiltration of inflammatory cells in the dermis. Psoriasis is also prone to induce a variety of complications, such as arthritis, metabolic syndrome, cardiovascular disease, mental illness, etc., which seriously affect the quality of life and survival cycle of patients.

[0003] CC Chemokine Ligand 17 (CCL17), also known as Thymic and Activation-Regulated Chemokine (TARC), is an important cellular chemokine. This chemokine belongs to the CC chemokine family and plays multiple roles in immune regulation and neuroregulation. CCL17 exerts chemotactic effects by binding to chemokine receptor 4 (CCR4) on the cell surface, thereby chemotacticizing the migration and recruitment of Th2 cells, skin T cells and regulatory T cells. The abnormal function of CCL17 is closely related to the occurrence and development of many diseases, such as tumors, cardiovascular diseases and allergic diseases. Therefore, CCL17 is regarded as one of the potential targets for drug development and is expected to provide new ideas and strategies for disease treatment and drug design. Summary of the invention

[0004] The purpose of the present invention is to address the deficiencies of the prior art and provide an application of CCL17 as a target in the preparation of a drug for preventing and treating psoriasis.

[0005] In order to solve the above technical problems, the technical solution adopted by the present invention is as follows: Use of CCL17 as a target in the development, screening and / or preparation of drugs for preventing and treating psoriasis.

[0006] Use of a CCL17 inhibitor in the preparation of a drug for preventing and / or treating psoriasis.

[0007] Furthermore, the CCL17 inhibitor is at least one of a monoclonal antibody, a small molecule compound or a nucleic acid inhibitor that specifically inhibits the activity of CCL17.

[0008] Furthermore, the medicine also includes pharmaceutically acceptable excipients.

[0009] Furthermore, the dosage form of the drug includes any one of injection, patch, powder, tablet, capsule, gel or ointment.

[0010] Furthermore, the administration method of the drug includes but is not limited to injection.

[0011] Application of CCL17 detection reagent in the preparation of psoriasis detection, diagnosis and / or prognosis evaluation products.

[0012] Furthermore, the product may be a reagent, a kit, a device or a system.

[0013] Furthermore, the CCL17 detection reagent is a reagent for detecting CCL17 activity by ELISA detection, Dot blot detection, Western blot detection or immunohistochemistry.

[0014] The drug can treat imiquimod-induced psoriasis in mice, alleviate the skin lesion symptoms and pathological manifestations of psoriasis mice, and reduce the expression level of inflammatory factors in the skin lesions. Therefore, the drug can alleviate the disease progression of psoriasis mice.

[0015] Compared with the prior art, the present invention has the following beneficial effects: The present invention proposes for the first time that CCL17 is used as a target for drugs to prevent and treat psoriasis. Experiments have shown that inhibiting CCL17 significantly improves imiquimod-induced psoriasis in mice, alleviates the symptoms and pathological manifestations of skin lesions in psoriasis mice, and reduces the expression level of inflammatory factors in skin lesions. The present invention confirms that targeting CCL17 can be used as an effective target for alleviating psoriasis, provides the application potential of targeting CCL17 in the treatment of psoriasis, and is of great significance for the prevention and treatment of psoriasis in the future. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 is a representative picture of the back skin of mice in each group; Figure 2 are the pathological representative hematoxylin-eosin (H&E) staining images of the skin lesions of mice in each group; Figure 3 The graphs show the expression levels of (A) IL-17A, (B) IL-22, (C) TNF-α, and (D) IL-10 in the skin lesions of mice in each group. DETAILED DESCRIPTION

[0017] The present invention is further described in detail below in conjunction with embodiments and drawings, but the embodiments of the present invention are not limited thereto.

[0018] The experimental methods described in the following examples are conventional methods unless otherwise specified, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. described in the following examples are all commercially available unless otherwise specified.

[0019] Example 1: Improvement of Imiquimod-induced Psoriasis in Mice by Inhibiting CCL17 1. Experimental Materials: Experimental animals: SPF BALB / C mice, weight: 20±2 g, age: 6 weeks, purchased from Guangdong Experimental Animal Center, male, free diet, temperature of 25±3℃, kept in SPF animal room.

[0020] CCL17 monoclonal antibody (clone number: 110904, catalog number: C1435) and isotype control antibody (clone number: 1-1, catalog number: I-1177) were purchased from Leinco Technologies.

[0021] 2. Grouping and Experiment Grouping: 25 BALB / C mice were randomly divided into 5 groups, 5 mice in each group, including normal group, model group, CCL17 monoclonal antibody low-dose group, CCL17 monoclonal antibody medium-dose group and CCL17 monoclonal antibody high-dose group.

[0022] Experiment: The backs of mice in each group were depilated with electric clippers and depilatory cream to form an exposed area of ​​about 2 cm × 2 cm. 62.5 mg of imiquimod ointment was applied to the depilated area on the back of the mice for 7 consecutive days to establish a mouse psoriasis model, and the normal group was applied with an equal amount of vaseline. Starting from the first day of applying imiquimod ointment, mice in the high, medium and low dose groups of CCL17 monoclonal antibody were intraperitoneally injected with 0.5, 1 and 2 μg of CCL17 monoclonal antibody every other day, and mice in the normal group and model group were intraperitoneally injected with 2 μg of isotype control antibody at the same time point. After the end, the back skin of the mice was taken for pathological observation and inflammatory factor detection.

[0023] 3. Methodological testing: (1) Changes in mouse skin appearance The daily changes in the skin of mice in each group were observed and recorded using a digital camera.

[0024] (2) Mouse skin pathology analysis Mouse skin specimens were fixed in 4% neutral formaldehyde, soaked overnight, dehydrated in a program dehydrator, embedded in a paraffin embedding machine, and then the paraffin-embedded tissue was cut into 4 μm thick sections. The sections were placed in a 65°C oven for 2 h, dewaxed to water, stained with hematoxylin for 10 min, rinsed with tap water to turn blue, stained with hydrochloric acid eosin for 15 min, dehydrated to make it transparent, sealed with neutral gum, dried overnight, and observed the pathological changes of skin lesions under a microscope.

[0025] (3) Detection of inflammatory factors Weigh 25 mg of mouse skin specimen, cut into small pieces, add 250 μl RIPA lysis buffer (containing protease inhibitors), and homogenize. Centrifuge at 12000g for 15 minutes and take the supernatant. According to the instructions of the ELISA kit, measure the expression levels of IL-17A, IL-22, TNF-α and IL-10.

[0026] 4. Experimental results: (1) Compared with the normal group, the mice in the model group had skin lesions on their backs, including thickening and scaling. After treatment with CCL17 monoclonal antibodies, the above symptoms were significantly improved. Moreover, the higher the treatment dose, the better the treatment effect. Figure 1 ).

[0027] (2) In terms of pathological histology, compared with the normal group, the back skin of the mice in the model group showed symptoms such as incomplete keratinization of the epidermis, thickening of the stratum spinosum, hyperplasia of the epidermal basal layer, and increased infiltration of inflammatory cells in the dermis, showing a psoriasis pathological state. After treatment with CCL17 monoclonal antibody, the above symptoms were significantly alleviated ( Figure 2 ).

[0028] (3) Inflammatory factor detection found that compared with the normal group, the expression levels of IL-17A, IL-22 and TNF-α in the back lesions of mice in the model group were significantly increased, while the expression level of IL-10 was not significantly different. When psoriasis mice were treated with CCL17 monoclonal antibody, the expression levels of IL-17A, IL-22 and TNF-α in their lesions were significantly reduced, and the expression level of IL-10 was significantly increased. The effect became more significant as the dose of CCL17 monoclonal antibody increased ( Figure 3 ).

[0029] The above results show that after inhibiting CCL17, the symptoms of scaling and thickening of the skin lesions in psoriasis mice were significantly alleviated, and the symptoms such as epidermal thickness and inflammatory cell infiltration in the dermis were significantly reduced. At the same time, the expression levels of IL-17A, IL-22 and TNF-α in the skin lesions were significantly reduced, and the expression level of IL-10 was significantly increased. The above results suggest that CCL17 plays an important role in the treatment of psoriasis.

[0030] Comparative Example 1: Effect of CCL17 knockout on imiquimod-induced psoriasis in mice 1. Experimental Materials: Experimental animals: SPF-grade C57BL / 6J mice and CCL17-KO mice (strain number: T028630, C57BL / 6J background), weight: 20±2 g, age: 6 weeks, purchased from Jiangsu Jicui Pharmaceutical Biotechnology Co., Ltd., free diet, temperature of 25±3℃, kept in SPF-grade animal room.

[0031] 2. Grouping and Experiment Grouping: The mice were divided into Wild type (WT) group and CCL17-KO group, with 6 mice in each group, half of which were male and half were female.

[0032] Experiment: The backs of mice in each group were depilated with electric clippers and depilatory cream to form an exposed area of ​​about 2 cm × 2 cm. 62.5 mg of imiquimod ointment was applied to the depilated area on the backs of mice for 7 consecutive days to establish a mouse psoriasis model. After the end of the study, the back skin of the mice was taken for pathological observation.

[0033] 3. Methodological testing: Mouse skin pathological analysis: Mouse skin specimens were fixed in 4% neutral formaldehyde, soaked overnight, dehydrated in a program dehydrator, embedded in a paraffin embedding machine, and then the paraffin-embedded tissue was cut into 4 μm thick sections. The sections were placed in a 65°C oven for 2 h, dewaxed to water, stained with hematoxylin for 10 min, rinsed with tap water to turn blue, stained with hydrochloric acid eosin for 15 min, dehydrated and transparent, sealed with neutral gum, dried overnight, observed under a microscope, photographed, and analyzed with ImagePro Plus 6.0 software for the thickness of the skin epidermis and the papilloma index. The thickness of the epidermis is the vertical distance between the upper epidermis and the lower epidermis, and the papilloma index = the length of the lower epidermis / the length of the upper epidermis.

[0034] 4. Experimental results: Pathological histological results showed that compared with the WT group, there was no significant decrease in the epidermal thickness of the back skin of mice in the CCL17-KO group, and there was no significant difference in the papilloma-like index.

[0035] Table 1 Epidermal thickness and papilloma index in mice The above results show that even directly knocking out the CCL17 gene cannot significantly alleviate the symptoms of psoriasis in mice. This is because knocking out CCL17 may cause immune compensation in mice, leading to the occurrence of psoriasis.

[0036] The present invention provides a concept and method for preparing a drug for preventing and treating psoriasis using CCL17 as a target. There are many methods and approaches to implement the technical solution. The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. Use of CCL17 as a target in the development, screening and / or preparation of drugs for the prevention and treatment of psoriasis.

2. Use of CCL17 inhibitors in the preparation of drugs for preventing and / or treating psoriasis.

3. The use according to claim 2, characterized in that: The CCL17 inhibitor is at least one of a monoclonal antibody, a small molecule compound or a nucleic acid inhibitor that specifically inhibits the activity of CCL17.

4. The use according to claim 2, characterized in that: The CCL17 inhibitor is a monoclonal antibody that specifically inhibits the activity of CCL17.

5. The use according to any one of claims 2 to 4, characterized in that: The medicine also includes pharmaceutically acceptable excipients.

6. The use according to any one of claims 2 to 4, characterized in that: The dosage form of the drug includes any one of injection, patch, powder, tablet, capsule, gel or ointment.

7. The use according to any one of claims 2 to 4, characterized in that: The administration method of the drug includes injection.

8. Use of CCL17 detection reagents in the preparation of psoriasis detection, diagnosis and / or prognosis assessment products.

9. The use according to claim 8, characterized in that: The product is a reagent, a kit, a device or a system.

10. The use according to claim 8 or 9, characterized in that: The CCL17 detection reagent is a reagent for detecting CCL17 activity through ELISA detection, Dot blot detection, Western blot detection or immunohistochemistry.

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