Application of ML348 in prevention and treatment of androgenetic alopecia

Abstract

The invention relates to the technical field of medicines, and discloses application of ML348 in prevention and treatment of androgenetic alopecia. In order to solve the technical problem of preventing and treating androgenetic alopecia, the ML348 is applied to intervention of DHT-induced AGA model mice, which shows that the ML348 has the effects of promoting hair regeneration, increasing hair length and generating hair follicle melanin, and can effectively resist hair loss caused by DHT. On the basis, a new scheme for treating the androgenetic alopecia is provided for preventing and / or treating the androgenetic alopecia.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to application of ML348 in preventing and treating androgenic alopecia. Background Art

[0002] Androgenetic Alopecia (AGA) is one of the most common hair loss diseases in the world. Although it does not affect the patient's physical health, it has a great negative impact on the patient's mental health and social identity, leading to a decrease in the patient's quality of life. AGA can occur in both men and women. Men usually show a significant receding hairline on the forehead and temples on both sides, and sparse hair on the top of the head; women show sparse and thin hair between the top of the head and the hairline, and the forehead hairline is retained. The pathogenesis of AGA is complex and is the result of the combined action of multiple factors, including genetics, androgen metabolism, spirit, diet, and stress. It is generally believed that androgens play an important role in the pathogenesis of AGA. Testosterone in the human body is converted into dihydrotestosterone (DHT) under the catalysis of 5α-reductase (SRD5A2). Studies have found that the activity of 5α-reductase in the scalp of AGA patients with hair loss is significantly higher than that in non-hair loss areas, and the DHT content is significantly increased in the hair loss area. Excessive DHT binds to the androgen receptor (AR), initiating signals that regulate hair follicle growth, causing hair follicle atrophy, a reduction in hair diameter, and a transformation of hair follicles from terminal hair to vellus hair. This shortens the growth period, affects the normal physiological cycle of hair, and ultimately leads to androgenic alopecia.

[0003] As AGA becomes more common, treatments for AGA are being developed. The main treatments for AGA include topical minoxidil, oral finasteride or spironolactone, hair transplant surgery, and LDL laser irradiation. The two most effective drugs for treating AGA are topical minoxidil and oral finasteride. Minoxidil increases vascular endothelial growth factor (VEGF) in dermal papilla cells in a dose-dependent manner and stimulates the production of prostaglandin E2, resulting in a prolonged growth phase. Side effects include contact dermatitis, headaches, and hirsutism. Short-term telogen shedding can be seen in the first 8 weeks of treatment. Finasteride is a class of drugs known as 5-alpha reductase inhibitors. These drugs block the 5-alpha reductase enzyme, thereby blocking the conversion of testosterone to the active form of dihydrotestosterone (DHT), slowing or reversing the hair loss process, but the use of finasteride may be accompanied by side effects such as sexual dysfunction. Non-drug treatments such as PRP and microneedling are good treatment options, but they are expensive and may not help patients with severe AGA. Hair transplantation is an excellent option for individuals who have failed medical treatment. There is not enough evidence to support the efficacy of other commonly used medications for this condition, such as topical and hormonal medications, and over-the-counter supplements. Additionally, the multifactorial origin and chronic nature of AGA make it difficult to treat. Therefore, there is an urgent need for more safe and effective treatment options.

[0004] AGA causes a serious psychological burden on the patient's body and mind. In the long run, it leads to mental disorders in patients, and in severe cases, depression and other symptoms. Healthy and beautiful hair makes people more confident in the process of communication, so people's demand for hair loss treatment is becoming more and more urgent. In this field, it is urgent to find more efficient and safe drugs for the prevention and treatment of androgenic alopecia to meet clinical needs. Summary of the invention

[0005] In order to solve the technical problem of androgenic alopecia, the present invention provides the use of ML348 in preventing and treating androgenic alopecia. The researchers of the present invention applied ML348 to DHT-induced AGA model mice and found that ML348 has the effects of promoting hair regeneration, increasing hair length and melanin production in hair follicles, and can effectively resist hair loss caused by DHT.

[0006] The specific technical scheme of the present invention is: In a first aspect, the present invention provides the use of ML348 in preparing a product for preventing and / or treating androgenic alopecia.

[0007] ML348 (CAS: 899713-86-1) is a depalmitoylation inhibitor that increases palmitoylation. Currently, it is commonly used in the treatment of renal fibrosis and Huntington's disease. The researchers of the present invention applied ML348 to intervene in DHT-induced AGA model mice and found that ML348 has the effects of promoting hair regeneration, increasing hair length and melanin production in hair follicles, and can effectively resist hair loss caused by DHT. Based on this result, the present invention can provide a new method for preventing and / or treating androgenic alopecia.

[0008] In their study of AGA patients, the researchers found that the expression of lysophospholipase 1 (LYPLA1) in the lesions of AGA patients was higher than that in the non-lesions. Multiple experiments have shown that this phenomenon may be related to the progression of AGA. Based on this result, the researchers further applied LYPLA1 The inhibitor ML348 intervened in DHT-induced AGA model mice and clarified the effect of ML348 on AGA intervention, hoping to provide a new solution for the prevention and treatment of AGA.

[0009] As a preferred embodiment of the above technical solution of the present invention, the product is a medicine, daily chemical product, food or food additive.

[0010] As a preferred embodiment of the above technical solution of the present invention, the product intervenes and / or treats androgenic alopecia by resisting the inhibition of hair follicle growth caused by DHT. The product is preferably a medicine, daily chemical product, food, or food additive that resists the inhibition of hair follicle growth caused by DHT.

[0011] The present invention explores the molecular mechanism and application of ML348 in alleviating DHT-induced androgenic alopecia in mice, clarifies the effectiveness of ML348 in inhibiting abnormal hair follicle growth in mice caused by DHT, and also proves the feasibility of ML348 in promoting hair regeneration, providing a theoretical basis for intervening and / or treating androgenic alopecia, and showing the feasibility of ML348 in intervening and / or treating androgenic alopecia by resisting the inhibition of hair follicle growth caused by DHT.

[0012] In a second aspect, the present invention provides a preparation for treating androgenic alopecia, wherein the active ingredient of the preparation includes ML348.

[0013] The researchers of the present invention applied ML348 to intervene in DHT-induced AGA mice, and confirmed that ML348 has the effects of promoting hair regeneration, increasing hair length and melanin production in hair follicles, and can effectively resist hair loss caused by DHT. Based on this, an effective biological product can be provided for preventing and / or treating androgenic alopecia, and the active ingredient of the product includes ML348.

[0014] As a preferred embodiment of the above technical solution of the present invention, the active ingredient of the product is ML348.

[0015] As a preferred embodiment of the above technical solution of the present invention, the product is a medicine, a daily chemical product, a food, or a food additive.

[0016] As a preferred embodiment of the above technical solution of the present invention, the product intervenes and / or treats androgenic alopecia by resisting the inhibition of hair follicle growth caused by DHT. The product is preferably a medicine, daily chemical product, food, or food additive that resists the inhibition of hair follicle growth caused by DHT. In addition to being used in the preparation of medicines, ML348 can also be used in the preparation of hair loss care products and skin external additives.

[0017] The present invention explores the application of ML348 in alleviating hair loss in mice under DHT stimulation and the related molecular mechanisms, clarifies the effectiveness of ML348 in inhibiting abnormal hair follicle growth in mice caused by DHT, and also proves the feasibility of ML348 in promoting hair regeneration, providing a basis for intervening or treating androgenic alopecia, and showing the feasibility of ML348 in intervening and / or treating androgenic alopecia by resisting the inhibition of hair follicle growth caused by DHT.

[0018] Compared with the prior art, the present invention has the following technical effects: The researchers of the present invention applied ML348 to intervene in DHT-induced AGA model mice, indicating that ML348 has the effects of promoting hair regeneration, increasing hair length and melanin production in hair follicles, and can effectively resist hair loss caused by DHT. Based on this, a new solution for the treatment of androgenic alopecia is provided for the prevention and / or treatment of androgenic alopecia. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 This is a diagram of hair growth of mice at different times in Example 1 of the present invention; Figure 2 This is a picture of H&E staining of the back skin of mice in each group after 21 days of treatment in Example 2 of the present invention; Figure 3 This is a diagram showing the melanin staining results of the mouse skin in each group in Example 3 of the present invention. DETAILED DESCRIPTION

[0020] The present invention will be further described below in conjunction with embodiments. Those of ordinary skill in the art will be able to implement the present invention based on these descriptions. In addition, the embodiments of the present invention involved in the following description are generally only embodiments of a part of the present invention, rather than all embodiments. Therefore, based on the embodiments in the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative work should belong to the scope of protection of the present invention.

[0021] The experimental animals used in the examples of the present invention were: 20 6-week-old male C57BL / 6 mice (25 g ± 0.5 g) purchased from the Experimental Animal Center of Hangzhou Medical College. All mice were raised under clean-grade conditions in the center.

[0022] In the embodiments of the present invention, the hematoxylin and eosin staining kit was purchased from Shanghai Biyuntian Biotechnology Co., Ltd.; dihydrotestosterone (methandrolone) was purchased from Shanghai McLean Biochemical Technology Co., Ltd.; ML348 was purchased from MCE; dimethyl sulfoxide (DMSO) was purchased from Sigma-Aldrich; and melanin staining solution (ferrous sulfate method) was purchased from Zhuhai Beso Biotechnology Co., Ltd.

[0023] Example 1 Establishment and drug administration of AGA mouse model (1) Experimental preparation Before the experiment, newly purchased male C57BL / 6 mice were fed for 7 days and then randomly divided into 4 experimental groups, with 5 mice in each group.

[0024] The four experimental groups are: Control group: only shaving was performed without any treatment.

[0025] DHT group: DHT was applied to the hair removal area on the back to induce the AGA model.

[0026] DHT / DMSO group: DHT was applied to the hair removal area on the back and DMSO was injected subcutaneously.

[0027] DHT / ML348 group: DHT was applied to the hair removal area on the back, supplemented with subcutaneous injection of ML348.

[0028] (2) Model establishment and drug administration Day 0: On the first day of the experiment, the backs of all mice were shaved and depilatory cream was applied to remove 2 square centimeters (2 cm × 2 cm) of hair.

[0029] Day 1~ Day 21: Starting from the first day of the experiment (Day 1), for 21 consecutive days, mice in the DHT group, DHT / DMSO group, and DHT / ML348 group were topically applied with 200 μL of DHT solution on their backs every day. At the same time, mice in the DHT / DMSO group received an additional subcutaneous injection of the same dose (200 μL) of DMSO solution every day; mice in the DHT / ML348 group received an additional subcutaneous injection of the same dose (200 μL) of ML348 solution every day.

[0030] (3) Observation and recording Photographing records: On the 7th, 14th and 21st days of the experiment, photographs were taken of the treated areas on the backs of the mice to record the growth of hair, such as Figure 1 As shown. Figure 1 shown. Figure 1 In the table, Control indicates the control group, DHT indicates the DHT group, DHT / DMSO indicates the DHT / DMSO group, and DHT / ML348 indicates the DHT / ML348 group.

[0031] Day 21: At the end of the experiment, back skin samples of all mice were collected for analysis of biological indicators in Examples 2 and 3.

[0032] Example 2 Hematoxylin-eosin (H&E) staining of mouse skin tissue A 1 cm2 (1 cm × 1 cm) skin sample was taken from the back of the mouse, fixed with 4% paraformaldehyde, dehydrated and embedded in paraffin, cut into 5 μm slices and baked in a 60°C oven. Then, the slices were treated in xylene for 15 min to remove the paraffin, and then immersed twice to ensure sufficient dewaxing. Next, the slices were placed in a hematoxylin aqueous solution for staining for 5 min, and then rinsed with running water for 15 min. After staining, the slices were dehydrated in 75% and 90% ethanol step by step, each for 10 min. Finally, the slices were immersed in 0.5% ethanol-eosin staining solution for 2 min and rinsed with running water for 1 min. After the stained slices were dehydrated with anhydrous ethanol, they were transparentized in xylene again for 3 min, and repeated once. After that, they were sealed with neutral resin. The sealed slices were placed under a microscope for observation and photography.

[0033] The results of H&E staining of the back skin of mice in each group after 21 days of treatment are as follows: Figure 2 As shown, including H&E staining observation results (a), hair follicle number statistics (b) and skin thickness statistics (c). Figure 2 In the table, Control indicates the control group, DHT indicates the DHT group, DHT / DMSO indicates the DHT / DMSO group, and DHT / ML348 indicates the DHT / ML348 group.

[0034] Example 3 Melanin staining of mouse skin Slice preparation: Place the slices of Example 2 in a 55°C constant temperature box for preheating for 30 minutes to complete dewaxing and antigen repair. Washing: Wash the slices once with distilled water. Melanin staining: Treat the slices with ferrous sulfate solution for 1 hour. Washing: Wash again with distilled water 3 times. Reaction: Add potassium ferrocyanide acetic acid solution and treat for 30 minutes. Washing: Wash with glacial acetic acid solution for 3 seconds and shake dry. VanGieson staining: Stain for 1 minute. Dehydration and transparency: First treat with 95% ethanol for 30 seconds, then dehydrate with anhydrous ethanol, and finally transparentize with xylene. Sealing: Sealing with neutral gum.

[0035] The melanin staining results of the mouse skin in each group were observed and photographed, and the melanin staining was quantitatively analyzed as follows: the IHC Toolbox of Image J software was used to select the color of the positive area, and then the melanin-stained positive area was automatically outlined, and the appropriate color threshold was adjusted. The image was adjusted to 8-bit format, and the threshold was adjusted to select the positive area and the entire stained area, and the area value was obtained. The percentage of the melanin-stained positive area / the total stained area was calculated to obtain the result.

[0036] Melanin staining results of mouse skin in each group are shown in Figure 3 . Figure 3 In the table, Control indicates the control group, DHT indicates the DHT group, DHT / DMSO indicates the DHT / DMSO group, and DHT / ML348 indicates the DHT / ML348 group.

[0037] Data analysis Figure 1 The back hair growth of mice in the control group, DHT group, DHT / DMSO group and DHT / ML348 group on days 0, 7, 14 and 21 of the experiment is shown. On the 14th day of the early experiment, the mice in the control group and DHT / ML348 group showed obvious hair regeneration, while the back skin of mice in the DHT group and DHT / DMSO group remained pink and no hair grew. The hair regeneration of mice in the DHT / ML348 group increased significantly, and the hair on the back grew a lot, showing a significant hair recovery effect. In contrast, the mice in the DHT group and DHT / DMSO group did not begin to show local hair regeneration until the 21st day of the experiment, but the hair growth was sparse and limited in area, far less than the hair coverage of the control group and DHT / ML348 group. The results show that ML348 has a significant effect on resisting the DHT-induced hair loss model and promoting hair regeneration.

[0038] Figure 2 H&E staining results of each treatment group are shown. Figure 2The staining results of a and 2b showed that the hair in the growth phase of the DHT / ML348 group was longer than that of the DHT group and the DHT / DMSO group, and the difference was statistically significant ( P <0.005). Meanwhile, the degree of hair regeneration in the DHT / ML348 group was similar to that in the Control group, indicating the potential of ML348 in promoting hair regeneration. Figure 2 a and 2c) showed that the dermis thickness of the Control group and the DHT / ML348 group was thicker than that of the DHT group and the DHT / DMSO group, and the difference was statistically significant ( P <0.05), but there was no statistically significant difference in dermal thickness between the Control group and the DHT / ML348 group. This suggests that ML348 can intervene in the inhibition of hair growth and thinning of dermal thickness caused by DHT, and has the effect of promoting hair regeneration.

[0039] Figure 3 The ferrous sulfate method was used to assess the skin melanin levels in different mouse groups. Figure 3 It can be seen that there is very little melanin staining in the skin of mice in the DHT group and the DHT / DMSO group. The melanin levels in the Control group and the DHT / ML348 group are higher than those in the DHT group and the difference is statistically significant (P<0.05), but there is no statistically significant difference between the Control group and the DHT / ML348 group. This shows that ML348 can promote melanin production.

[0040] Therefore, based on the above results, it is shown that ML348 has the effect of promoting hair regeneration, increasing hair length and melanin production in hair follicles. It can effectively resist hair loss caused by DHT, showing the potential value of ML348 in the treatment of AGA and providing a new solution for the treatment of AGA.

[0041] The raw materials and equipment used in the present invention, unless otherwise specified, are all commonly used raw materials and equipment in the art; the methods used in the present invention, unless otherwise specified, are all conventional methods in the art.

[0042] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any way. Any simple modification, change and equivalent transformation made to the above embodiment based on the technical essence of the present invention still falls within the protection scope of the technical solution of the present invention.

Claims

1. Use of ML348 in the preparation of products for preventing and / or treating androgenic alopecia.

2. The use according to claim 1, characterized in that: The product is medicine, daily chemical product, food, or food additive.

3. The use according to claim 2, characterized in that: The product is a medicine, daily chemical product, food, or food additive for resisting the inhibition of hair follicle growth caused by DHT.

4. A product for treating androgenic alopecia, characterized in that: The active ingredient of the preparation includes ML348.

5. A product for treating androgenic alopecia according to claim 4, characterized in that: The active ingredient of the preparation is ML348.

6. A product for treating androgenic alopecia according to claim 4, characterized in that: The product is medicine, daily chemical product, food, or food additive.

7. A product for treating androgenic alopecia according to claim 6, characterized in that: The product is a medicine, daily chemical product, food, or food additive for resisting the hair follicle growth inhibition caused by DHT.

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