Compound for treating androgen alopecia, preparation method thereof and application of compound as SFRP1 (secreted frizzled-related protein 1) inhibitor

Abstract

The invention discloses a compound for treating androgen alopecia, a preparation method thereof and application of the compound as an SFRP1 inhibitor, and relates to the technical field of pharmaceutical preparations. The compound provided by the invention can target SFRP1 protein in a Wnt / beta-catenin pathway and generate strong affinity with the SFRP1 protein, so that the Wnt / beta-catenin signaling pathway is activated, hair follicle cell growth is further promoted, hair growth is promoted, skin irritation is small, tolerance is good, and the effect of effectively treating androgenetic alopecia is achieved.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical formulation technology, and in particular to a compound for treating androgenetic alopecia, its preparation method, and its application as an SFRP1 inhibitor. Background Technology

[0002] Androgenetic alopecia (AGA) is the most common type of hair loss worldwide. It is characterized by progressive miniaturization of hair follicles and shortening of the growth phase, eventually leading to the gradual replacement of terminal hair (thick, dark, with a long growth phase) with vellus hair (fine, soft, with a short growth phase), resulting in sparse hair on the scalp.

[0003] Androgenetic alopecia is increasingly affecting younger people, and its mechanisms involve the interaction of multiple factors, including genetics, androgens, and the hair follicle microenvironment. Currently, the FDA-approved drugs for treating androgenetic alopecia are only 5% minoxidil and 1 mg finasteride. While these drugs have shown varying degrees of efficacy, they also have varying degrees of side effects. For example, minoxidil can cause scalp irritation, itching, and excessive hair growth on the face or body (hirsutism), and discontinuing the drug can lead to renewed hair loss. Men who take finasteride long-term may experience side effects such as erectile dysfunction, decreased libido, and gynecomastia, which may persist even after treatment is stopped.

[0004] In recent years, drug development research for androgenetic alopecia (AGA) has primarily focused on overcoming the limitations of traditional drugs and developing novel targeted therapies at the molecular mechanism analysis and treatment strategy exploration levels. Specifically, current research directions encompass androgen receptor inhibitors, Wnt pathway agonists, thyroid hormone receptor modulators, prostaglandin-related modulators, PDE4 inhibitors, and RIPK1 inhibitors, providing new technological pathways to address the issues of "limited efficacy and significant side effects" in the treatment of androgenetic alopecia. These emerging research directions not only provide new perspectives for a deeper understanding of the complex pathological mechanisms of androgenetic alopecia but also lay the foundation for developing safer, more effective, and more targeted therapeutic drugs. In the future, it is hoped that this will break the 40-year drought of new drugs for treating androgenetic alopecia, bringing more ideal treatment options to patients and effectively improving their quality of life.

[0005] The Wnt signaling pathway exists in various organs of the human body and participates in basic biological processes such as cell proliferation and differentiation. The classic pathway, in which Wnt protein binds to its receptor, activates β-catenin to enter the cell nucleus, and initiates the transcription of target genes, is the Wnt / β-catenin pathway. In skin hair follicle tissue, Wnt signaling regulates the growth cycle of hair matrix cells. Once Wnt signaling is blocked, the hair follicle enters the anagen phase prematurely, leading to hair loss. Experiments have shown that Wnt signaling activation can drive hair follicles to restart the anagen phase from the telogen phase, promoting new hair growth. In short, the Wnt / β-catenin signaling pathway is the "master switch" regulating the hair cycle. Developing drugs that activate this pathway has become a new strategy for treating hair loss.

[0006] Secreted frizzled-related protein 1 (SFRP1) is a member of the SFRP family of secreted glycoproteins. As a primitive gene regulating cell growth, development, and transformation, SFRP1 is widely expressed in human cells. As a secreted glycoprotein, SFRP1 is considered a natural antagonist of the Wnt pathway due to its high structural homology with the frizzled (Fz) receptor for Wnt signaling. It can interfere with Wnt signal transduction and participate in the growth and apoptosis of hair follicle cells. Targeted inhibition of SFRP1 can activate the Wnt / β-catenin signaling pathway, further promoting hair follicle cell growth and hair growth, representing a new direction for the development of new drugs for treating hair loss. Compared to directly developing Wnt agonists, targeted inhibition of SFRP1 avoids the side effects caused by systemic overactivation of the pathway. Furthermore, as a single target, SFRP1's mechanism of action is relatively clear (inhibiting the pathway by competitively binding to Wnt with the frizzled receptor), making the development of specific inhibitors easier and enabling precise regulation.

[0007] In view of this, the present invention is proposed. Summary of the Invention

[0008] The purpose of this invention is to provide a compound for treating androgenetic alopecia, its preparation method, and its application as an SFRP1 inhibitor, thereby addressing the problems existing in the prior art. The compound of this invention can target the SFRP1 protein in the Wnt / β-catenin pathway, exhibiting a strong affinity for SFRP1, thereby activating the Wnt / β-catenin signaling pathway, further promoting hair follicle cell growth and hair growth. Its effects are significant, without skin irritation or side effects, achieving an effective treatment for androgenetic alopecia.

[0009] To achieve the above objectives, the present invention provides the following solution:

[0010] This invention provides a compound for treating androgenetic alopecia, having the structure shown in any one of formulas (I)-(IV):

[0011]

[0012] Wherein, R is one of methyl, alkoxy, trifluoromethyl, halogen, carbamate, sulfonamide, amino, and sulfone groups;

[0013] X and Y are independently selected from carbon or nitrogen atoms;

[0014] W represents either a carbon atom or a nitrogen atom.

[0015] The second technical solution of the present invention is to provide the above-mentioned compound for treating androgenetic alopecia as an SFRP1 specific inhibitor.

[0016] The third technical solution of the present invention is to provide the application of the above-mentioned compound for treating androgenetic alopecia in the preparation of a drug for treating androgenetic alopecia.

[0017] The fourth technical solution of the present invention provides a pharmaceutical composition for treating androgenetic alopecia, comprising a compound with the structure shown in any one of formulas (I) to (IV) as the active pharmaceutical ingredient.

[0018] As a further preferred embodiment of the present invention, the dosage form of the pharmaceutical composition includes aerosol, solution, gel, cream, ointment or film.

[0019] The Wnt signaling pathway exists in various organs of the human body and participates in basic biological processes such as cell proliferation and differentiation. The classic pathway, in which Wnt protein binds to its receptor, activates β-catenin to enter the cell nucleus, and initiates the transcription of target genes, is the Wnt / β-catenin pathway. In skin hair follicle tissue, Wnt signaling regulates the growth cycle of hair matrix cells. Once Wnt signaling is blocked, the hair follicle enters the anagen phase prematurely, leading to hair loss. Experiments have shown that Wnt signaling activation can drive hair follicles to restart the anagen phase from the telogen phase, promoting new hair growth. In short, the Wnt / β-catenin signaling pathway is the "master switch" regulating the hair cycle. Developing drugs that activate this pathway has become a new strategy for treating hair loss.

[0020] Secreted frizzled-related protein 1 (SFRP1) is a member of the SFRP family of secreted glycoproteins. As a primitive gene regulating cell growth, development, and transformation, SFRP1 is widely expressed in human cells. As a secreted glycoprotein, SFRP1 is considered a natural antagonist of the Wnt pathway due to its high structural homology with the frizzled (Fz) receptor for Wnt signaling. It can interfere with Wnt signal transduction and participate in the growth and apoptosis of hair follicle cells. Targeted inhibition of SFRP1 can activate the Wnt / β-catenin signaling pathway, further promoting hair follicle cell growth and hair growth, representing a new direction for the development of new drugs for treating hair loss. Compared to directly developing Wnt agonists, targeted inhibition of SFRP1 avoids the side effects caused by systemic overactivation of the pathway. Furthermore, as a single target, SFRP1's mechanism of action is relatively clear (inhibiting the pathway by competitively binding to Wnt with the frizzled receptor), making the development of specific inhibitors easier and enabling precise regulation. Therefore, this invention treats androgen-induced hair loss by preparing compounds that can target the SFRP1 protein.

[0021] The present invention discloses the following technical effects:

[0022] The compound provided by this invention can target the SFRP1 protein in the Wnt / β-catenin pathway, generate a strong affinity for the SFRP1 protein, thereby activating the Wnt / β-catenin signaling pathway, further promoting hair follicle cell growth, promoting hair growth, having low skin irritation and good tolerability, and achieving an effective treatment for androgenetic alopecia. Attached Figure Description

[0023] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0024] Figure 1 The figure shows the fitting results of the response signal as a function of the concentration of compound WH1.

[0025] Figure 2 The figure shows the fitting results of the response signal as a function of the concentration of compound WH4.

[0026] Figure 3 The figure shows the fitting results of the response signal as a function of the concentration of compound WH2.

[0027] Figure 4This is a graph showing the changes in body weight of mice in each group after drug administration.

[0028] Figure 5 This is a visual representation of hair growth on the backs of mice in each group after drug administration.

[0029] Figure 6 The images show the pathological results of HE staining of the dorsal skin of mice in each group (A: normal control group; B: model group; C: positive drug group; D: test drug group). Detailed Implementation

[0030] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0031] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0032] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0033] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be readily apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0034] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0035] It should be noted that any aspects not described in detail in this invention are conventional practices in the field and are not the focus of this invention.

[0036] Unless otherwise specified, room temperature in this invention refers to a temperature of 20±10℃.

[0037] I. Preparation of Compounds

[0038] The WH1-WH4 compounds have the general structure shown in formula (II), and the substituents are shown in Table 1:

[0039] Table 1

[0040]

[0041] The synthetic routes for the WH1-WH4 compounds are shown below:

[0042]

[0043] The preparation method steps are as follows:

[0044] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of pyridineboronic acid or phenylboronic acid with different substitutions into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0045] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0046] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours, and the crude product was purified by preparative liquid chromatography to obtain WH1-WH4.

[0047] The preparation of WH1-WH4 compounds is described in detail below with reference to examples:

[0048] Example 1: Preparation of compound WH1

[0049]

[0050] The preparation steps are as follows:

[0051] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 4-tert-butylphenylboronic acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0052] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0053] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH1 with a yield of 38.4%.

[0054] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ8.56(s,1H).7.33-7.46(m,6H),6.96(d,2H,J=8.0Hz),4.50(s,1H),3.60(m,8H),1.87(s,4H),1.30(s,9H).

[0055] Example 2 Preparation of compound WH2

[0056]

[0057] The preparation steps are as follows:

[0058] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 2-methoxyphenylboronic acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0059] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0060] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH2 with a yield of 13.2%.

[0061] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ8.50(s,1H),7.45(m,2H),7.26(m,2H),6.95(m,2H),6 .85(m,2H),4.55(s,1H),3.80(s,3H),3.67(m,6H),3.41(m,2H),1.93(s,4H).

[0062] Example 3: Preparation of compound WH3

[0063]

[0064] The preparation steps are as follows:

[0065] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 3-fluoropyridine-4-boric acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0066] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0067] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH3 with a yield of 35.0%.

[0068] The NMR characterization results are as follows: 1 H NMR (400MHz, MeOD): δ8.59(s,1H),8.46(s,1H),7.71(m,3H),7.16(s,2H,J=8.8Hz ),4.80(m,1H),3.78(s,4H),3.68(m,2H),3.45(m,2H),2.10(m,2H),1.91(m,2H).

[0069] Example 4: Preparation of compound WH4

[0070]

[0071] The preparation steps are as follows:

[0072] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 2-fluoropyridine-4-boric acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0073] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0074] (3) Intermediate 2 and 1 equivalent of 2-bromo-1,4,5,6-tetrahydropyrimidine were dissolved in anhydrous methanol. The reaction mixture was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH4 with a yield of 9.1%.

[0075] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ8.56(m,1H),8.15(s,1H),7.84(m,1H),7.35(m,1H),7.25(m,1H),7 .15(m,1H),7.05(m,1H),6.87(m,1H),4.58(m,1H),3.71(s,6H),3.60(s,2H),1.95(s,4H).

[0076] The WH5-WH8 compounds have the general structure shown in formula (I), and the substituents are shown in Table 2:

[0077] Table 2

[0078]

[0079] The synthetic routes for the WH5-WH8 compounds are shown below:

[0080]

[0081] The preparation method of WH5-WH8 compounds includes the following steps:

[0082] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of pyridineboronic acid or phenylboronic acid with different substitutions into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0083] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0084] (3) Intermediate 2 and 1 equivalent of 2-bromo-1,4,5,6-tetrahydropyrimidine were dissolved in anhydrous methanol. The reaction mixture was stirred at 70°C for 10 hours, and the crude product was purified by preparative liquid chromatography to obtain WH5-WH8.

[0085] The preparation of WH5-WH8 will be described in detail below with reference to the embodiments.

[0086] Example 5: Preparation of compound WH5

[0087]

[0088] The preparation steps are as follows:

[0089] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 4-tert-butylphenylboronic acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0090] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0091] (3) Intermediate 2 and 1 equivalent of 2-bromo-1,4,5,6-tetrahydropyrimidine were dissolved in anhydrous methanol. The reaction mixture was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH5 with a yield of 32.8%.

[0092] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ7.82(s,1H).7.23-7.46(m,6H),6.96(d,2H,J=8.8Hz), 4.50(s,1H),3.60(s,2H),3.47(s,2H),3.34(s,4H),1.87(s,6H),1.32(s,9H).

[0093] Example 6 Preparation of compound WH6

[0094]

[0095] The preparation steps are as follows:

[0096] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 2-methoxyphenylboronic acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0097] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0098] (3) Intermediate 2 and 1 equivalent of 2-bromo-1,4,5,6-tetrahydropyrimidine were dissolved in anhydrous methanol. The reaction mixture was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH6 with a yield of 28.3%.

[0099] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ7.45(m,2H),7.26(m,2H),6.95(m,2H),6.85(m,2H),4 .56(s,1H),3.82(m,3H),3.61(m,2H),3.48(m,2H),3.35(m,4H),1.93(s,6H).

[0100] Example 7 Preparation of compound WH7

[0101]

[0102] The preparation steps are as follows:

[0103] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 3-fluoropyridine-4-boric acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0104] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0105] (3) Intermediate 2 and 1 equivalent of 2-bromo-1,4,5,6-tetrahydropyrimidine were dissolved in anhydrous methanol. The reaction mixture was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH7 with a yield of 42.3%.

[0106] The NMR characterization results are as follows: 1 H NMR (400MHz, MeOD): δ8.59(s,1H),8.46(s,1H),7.71(m,3H),7.16(s,2H,J=8.8Hz ),4.80(m,1H),3.60(m,2H),3.41(m,6H),2.15(m,2H),2.10(m,2H),1.91(m,2H).

[0107] Example 8 Preparation of compound WH8

[0108]

[0109] The preparation steps are as follows:

[0110] (1) Weigh 1-BOC-4-(4-bromophenoxy)piperidine and 1 equivalent of 2-fluoropyridine-4-boric acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0111] (2) Intermediate 1 was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the solution was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0112] (3) Intermediate 2 and 1 equivalent of 2-bromo-1,4,5,6-tetrahydropyrimidine were dissolved in anhydrous methanol. The reaction mixture was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH8 with a yield of 67.5%.

[0113] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ8.15(s,1H),7.84(m,1H),7.35(m,1H),7.25(m,1H),7.15(m,1H),7 .05(m,1H),6.87(m,1H),4.58(m,1H),3.68(s,2H),3.50(s,2H),3.37(s,4H),1.95(s,6H).

[0114] The WH9-WH12 compounds have the general structure shown in formula (IV), and the substituents are shown in Table 3:

[0115] Table 3

[0116]

[0117] The synthetic routes for the WH9-WH12 compounds are shown below:

[0118]

[0119] The preparation method of WH9-WH12 compounds includes the following steps:

[0120] (1) Weigh 1-BOC-4-(3-bromophenoxy)piperidine and 1 equivalent of pyridineboronic acid or phenylboronic acid with different substitutions into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0121] (2) The intermediate was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the product was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0122] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide or 2-bromo-1,4,5,6-tetrahydropyrimidine were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours, and the crude product was purified by preparative liquid chromatography to obtain WH9-WH12.

[0123] The preparation of compounds WH9-WH12 is described in detail below with reference to specific examples:

[0124] Example 9 Preparation of compound WH9

[0125]

[0126] The preparation steps are as follows:

[0127] (1) Weigh 1-BOC-4-(3-bromophenoxy)piperidine and 1 equivalent of 4-tert-butylphenylboronic acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0128] (2) The intermediate was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the product was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0129] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH9 with a yield of 10.9%.

[0130] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ7.50(m,4H),7.45(m,1H),7.25(m,1H),7.10(m,1H),6 .86(m,1H),4.60(s,1H),3.75(s,6H),3.51(m,2H),2.00(m,4H),1.38(m,9H).

[0131] Example 10 Preparation of compound WH10

[0132]

[0133] The preparation steps are as follows:

[0134] (1) Weigh 1-BOC-4-(3-bromophenoxy)piperidine and 1 equivalent of 2-fluorophenylboronic acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0135] (2) The intermediate was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the product was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0136] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH10 with a yield of 24.2%.

[0137] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ8.50(s,1H),7.32(m,3H),6.96-7.10(m,4H),6.82(m,1H),4.60(m,1H),3.75-3.47(m,8H),2.00(s,4H).

[0138] Example 11 Preparation of compound WH11

[0139]

[0140] The preparation steps are as follows:

[0141] (1) Weigh 1-BOC-4-(3-bromophenoxy)piperidine and 1 equivalent of 3-fluoropyridine-4-boric acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0142] (2) The intermediate was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the product was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0143] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH11 with a yield of 21.4%.

[0144] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ8.16(s,1H),7.88(m,1H),7.36(m,2H),7.32(m,1H),7.16(m, 1H),7.06(m,1H),4.60(s,1H),3.75(s,4H),3.61(s,2H),3.51(s,2H),2.00(s,4H).

[0145] Example 12 Preparation of compound WH12

[0146]

[0147] The preparation steps are as follows:

[0148] (1) Weigh 1-BOC-4-(3-bromophenoxy)piperidine and 1 equivalent of 2-fluoropyridine-4-boric acid into a reaction flask, add 1.5 equivalents of Pd(dppf)Cl2 and 2 equivalents of potassium carbonate, and add a mixed solution of N,N-dimethylacetamide and water. Stir under nitrogen protection and reflux at 100℃. Monitor the reaction by TLC until completion. After filtering and concentrating the solution, extract with ethyl acetate, wash three times with water and saturated brine, and dry the organic layer with magnesium sulfate. The crude product concentrated under reduced pressure is purified by silica gel column chromatography with dichloromethane / methanol = 50:1 as the eluent to obtain intermediate 1.

[0149] (2) The intermediate was added to a methanol and hydrochloric acid solution and stirred overnight at room temperature. The reaction was detected by TLC and the product was concentrated to obtain intermediate 2. The crude product was not purified and proceeded directly to the next step.

[0150] (3) Intermediate 2 and 1 equivalent of 2-methylthio-2-imidazoline hydrogen iodide were dissolved in anhydrous methanol. The reaction solution was stirred at 70°C for 10 hours. The crude product was purified by preparative liquid chromatography to obtain a white solid WH12 with a yield of 21.4%.

[0151] The NMR characterization results are as follows: 1 H NMR (400MHz, CDCl3): δ8.16(s,1H),7.88(m,1H),7.36(m,2H),7.32(m,1H),7.16(m, 1H),7.06(m,1H),4.60(s,1H),3.75(s,4H),3.61(s,2H),3.51(s,2H),2.00(s,4H).

[0152] II. Preparation of pharmaceutical compositions containing WH1 compounds

[0153] 1. Preparation of gelling agents

[0154] The composition and content of the gelling agent are shown in Table 4:

[0155] Table 4 Composition and content of gelling agent

[0156]

[0157] Preparation method of gelling agent:

[0158] Weigh carbomer 980 into a beaker, add an appropriate amount of water, and heat in a water bath at 80°C to dissolve. Separately weigh compound WH1, dissolve it in an appropriate amount of water, add it to the carbomer 980 solution, and finally add triethanolamine solution. Stir until a gel-like consistency is formed to obtain the final product.

[0159] 2. Preparation of the solution:

[0160] The composition and content of the solution are shown in Table 5:

[0161] Table 5 Composition and content of the solution

[0162]

[0163] Solution preparation method:

[0164] Weigh out compound WH1, add water to 10g, stir to dissolve, and you have the product.

[0165] 3. Preparation of ointments:

[0166] The composition and content of the ointment are shown in Table 6:

[0167] Table 6 Composition and content of ointments

[0168]

[0169] Solution preparation method:

[0170] Weigh out octadecyl alcohol, white petrolatum, and liquid paraffin and place them in a beaker. Heat in a water bath to 70–80°C to melt. Separately weigh out compound WH1, sodium lauryl sulfate, ethylparaben, glycerin, and water and place them in a beaker. Heat in a water bath to 70–80°C to dissolve. Mix the two liquids together, stir, and let cool to obtain the final product.

[0171] 4. Preparation of the film-forming agent:

[0172] The composition and content of the film-forming agent are shown in Table 7:

[0173] Table 7 Composition and content of film-forming agents

[0174] Element Mass (g) content(%) effect WH1 0.5 5 Main drug Polyvinyl alcohol 17-88 2.5 9 Film-forming materials glycerin 0.5 0.5 Moisturizer water 7 70 Aqueous phase / solvent

[0175] Preparation method of film-forming agent:

[0176] Weigh 17-88g of polyvinyl alcohol and place it in a beaker. Add water and soak overnight. Dissolve in an 80°C water bath and cool until bubbles overflow. Separately weigh WH1 compound, glycerin, and an appropriate amount of water, dissolve them, and add them to the polyvinyl alcohol paste. Stir and let stand until bubbles completely overflow. Coat the paste on a glass plate and dry at room temperature.

[0177] III. Effect Verification

[0178] 1. Single concentration screening of WH1-WH12:

[0179] Protein fixation:

[0180] Step 1: Activate the chip by mixing 100 μL of NHS with 100 μL of EDC at a flow rate of 10 μL / min;

[0181] Step 2: The protein was diluted to 40 μg / mL with sodium acetate at pH 4.0 and then fixed at a flow rate of 10 μL / min;

[0182] Step 3: Seal the chip surface using Ethamolamine-HCl.

[0183] The protein fixation results are shown in Table 8.

[0184] Table 8 Protein fixation results

[0185] protein Fixed level SFRP1 5000RU

[0186] Experimental setup: The experimental instrument used was a Biacore T200 (GE Healthcare), and the sensor chip was a CM5 sensor chip (Cytiva Lot 10324487). Buffer solution: PBS 10X (1.37M NaCl; 26.8mM KCl; 81mM Na2HPO4; 17.6mM KH2PO4; pH 7.2-7.4) (Sangon Biotech (Shanghai) Lot I308FD0198).

[0187] pH Souting: Prepare four 1.5 mL EP tubes. Dilute the SFRP1 ligand stock solution to 20 μg / mL with 10 mM sodium acetate solution at pH values ​​of 4.0, 4.5, 5.0, and 5.5, respectively, to a final volume of 100 μL for each tube. Remove air bubbles by centrifugation. In another 1.5 mL EP tube, add 200 μL of 50 mM sodium hydroxide. Run the manual run (Run→Manual Run) to inject the sample and determine the optimal pH conditions.

[0188] Conjugation and immobilization of ligands: Based on the pH southing results, the protein SFRP-1 was diluted to 50 μg / mL with sodium acetate at pH 4.5. The automatic conjugation program (aiming for immobilized level) was run, with a target conjugation amount of 10500 RU and a final conjugation amount of 5000 RU. Setting up the experimental program: Click "LMW SCREEN" in the New Wizard Template to begin setting up the experimental program. See Table 9 for the selected experimental program settings.

[0189] Table 9 Screening Experiment Procedure Settings

[0190]

[0191] The response signal values ​​of compounds WH1-WH12 at a single concentration (10 μM) are shown in Table 10:

[0192] Table 10 Response signal values ​​of compounds WH1-WH12 at a single concentration (10 μM)

[0193] Compound ID Response value (RU) WH1 4.18 WH4 3.14 WH3 3.06 WH2 3.10 WH5 3.03 WH8 3.01 WH7 2.89 WH6 2.76 WH12 2.71 WH11 2.66 WH9 2.60 WH10 2.58

[0194] The response values ​​in the table show that compounds WH1-WH12 all exhibited good signal performance at a single concentration (10 μM), indicating that compounds WH1-WH12 can bind to SFRP1. Among them, compounds WH1, WH4, and WH2 showed the best signal performance, and therefore were selected for subsequent quantitative binding experiments.

[0195] 2. In vitro binding affinity tests of compounds WH1, WH4, and WH2:

[0196] Protein fixation and experimental procedures are the same as those in Section 1 for single-concentration screening experiments. The quantitative experimental procedure settings are shown in Table 11.

[0197] Table 11 Quantitative Experimental Procedure Settings

[0198]

[0199] The analytical results for the WH1 compound group are shown below. Figure 1 . Figure 1 The fitting results for the response signal as a function of the concentration of compound WH1 are shown. The analytical results for the compound WH4 group are shown below. Figure 2 . Figure 2 The results show the fitting of the response signal of compound WH4 as a function of concentration. The analytical results for the WH2 compound group are shown below. Figure 3 . Figure 3 The fitting results are for the response signal as a function of the concentration of compound WH2.

[0200] 3. Efficacy evaluation of in vivo treatment for androgenetic alopecia:

[0201] The in vitro binding affinity tests of compounds WH1, WH4, and WH2 all showed stable-state fits, with KD values ​​(M) of 79.1 nM, 87.2 nM, and 252 nM, respectively. The affinity ranking was WH1 > WH4 > WH2, indicating strong binding between proteins and small molecules in the in vitro interaction domain. Subsequent experiments selected compound WH1, with the strongest binding affinity, as a lead compound for animal validation.

[0202] Test system:

[0203] Information on the experimental animals is shown in Table 12:

[0204] Table 12 Information on experimental animals

[0205] content Detailed information Species / Strain C57 / BL / 6 mice grade SPF gender male quantity 20 Zhou Ling 7 weeks source Jiangsu Huachuang Xinno Pharmaceutical Technology Co., Ltd. Animal Certificate Number B202505280226 Animal testing ethics HH-DWLL-YW2025007-1

[0206] Animal facilities:

[0207] Temperature / Humidity: Controlled within the following ranges: Temperature: (23±3)℃, Humidity: 40-70%

[0208] Cages: Made of polycarbonate. Dimensions: 370mm x 155mm x 135mm. Soft corncob pressure-sterilized bedding, changed twice weekly. Each cage has a tag indicating the number of animals, sex, strain, arrival time, group, and trial start time.

[0209] Feed, bedding, and water: The rats and mice were fed a diet suitable for growth and reproduction. The bedding consisted of corn cobs, and the drinking water was produced by a special experimental animal water dispenser. The animals had free access to food and water.

[0210] Animal ID: Ear piercing.

[0211] Animal experiment design:

[0212] The experimental grouping and dosing information are shown in Table 13.

[0213] Table 13 Trial Groups and Drug Administration

[0214]

[0215] Note: *Administration method: Spray once a day on the hair removal area of ​​the back, massage with fingertips until absorbed; #Administer according to the instructions.

[0216] Test method:

[0217] Preparation of test sample and reference standard:

[0218] Preparation of 5 mg / mL testosterone propionate injection: Each time, draw 1 mL (25 mg / mL) of testosterone propionate injection stock solution, add 4 mL of soybean oil, mix gently, avoid vigorous shaking to prevent air bubbles, protect from light, and seal and store at 2-8℃; during modeling, inject 0.02 mL subcutaneously at multiple points on the back of each mouse.

[0219] Preparation of 5% WH1 compound drug formulation: Dissolve 0.5g of WH1 powder, 0.02g of ethylparaben, and 0.03g of 2,6-tert-butyl-p-cresol in 10mL of distilled water. Store in a small spray bottle at 4°C protected from light.

[0220] Preparation of androgenetic alopecia model:

[0221] Fifteen mice were selected, and a 2cm x 4cm area on their backs was selected as the hair removal area. Hair removal cream was used to remove the hair, and multiple subcutaneous injections of 5mg / mL testosterone propionate solution were given to the backs. The injection volume was 0.02mL per mouse, once a day for 3 consecutive weeks. The hair removal area of ​​the mice was photographed weekly.

[0222] Grouping and administration:

[0223] Before grouping, 15 model mice were randomly divided into 3 groups according to their weight after hair removal cream removal: model group, positive drug group, and test drug group. Normal mice were selected as normal control group. Each group consisted of 5 mice. The day of grouping and drug administration was recorded as D0. Grouping and drug administration information are shown in Table 13.

[0224] Testing indicators:

[0225] Weight measurement and general condition observation: The weight of mice in each group was measured twice a week after drug treatment, and the health status of the animals was monitored. The hair loss area of ​​each group of mice was photographed weekly.

[0226] Hair length and weight determination: On days 14 and 20, five hairs were collected from the middle of the hairless area of ​​each animal, and the hair length was measured with calipers. On day 20, the mice were euthanized, and circular skin patches were taken from the same area of ​​the hairless area on the back of each mouse using a 10mm punch. All the hair on the skin patches was scraped off with a scalpel, and the hair weight was measured.

[0227] HE staining (1 mouse per group): At the end of the experiment, mice in each group were euthanized, and skin from the hairless area on the back was taken for HE staining and pathological analysis.

[0228] Experimental results:

[0229] mouse weight

[0230] During the administration period, at D19, one mouse in the model group (G2) died. The body weight of the dead mouse did not decrease significantly, and no abnormalities were found in the major organs upon gross dissection. The cause may be that the continuous administration of androgens caused the accidental death of the mouse. The remaining mice in the group did not decrease significantly in body weight and no other abnormalities were observed. The mice in the other groups were in good general condition, with no significant decrease in body weight and no other abnormalities were observed.

[0231] Androgenetic alopecia model was established by subcutaneously injecting testosterone propionate at multiple points on the back of C57BL / 6J mice. The model mice were randomly divided into three groups according to body weight: a model group (G2), a positive control group (G3, minoxidil, qd×20), a WH1 compound group (G4, 5% WH1, qd×20), and a normal control group (n=5 per group). Weekly photographs of the bald areas were taken. On days 14 and 20, five hairs were collected from the center of the bald area of ​​each animal, and their length was measured with calipers. On day 20, the mice were euthanized, and circular skin flaps were taken from the same location on the back of each mouse using a 10mm punch. All hair on the skin flaps was scraped off with a scalpel, and the hair weight was recorded. The mouse skin was fixed and stained with hematoxylin and eosin (HE). The therapeutic effect of the drug on androgenetic alopecia was evaluated by pathological HE staining.

[0232] Mouse hair length

[0233] At D14 and D20, the length of newly grown hair in the model group mice was significantly lower than that in the normal control group (p<0.01), indicating that the androgenic alopecia model was successful.

[0234] At days 14 and 20, compared with the model group, the length of newly grown hair in mice in the positive control (minoxidil) group and the test drug (WH1) group was significantly increased (p < 0.01). Notably, at days 14 and 20, the length of newly grown hair in the test drug (WH1) group was greater than that in the positive control (minoxidil) group, but the difference was not statistically significant. This is likely due to the smaller number of mice in each group. The results are shown in Table 14 and... Figure 5 .

[0235] Table 14 Hair length of mice in each group after drug administration (mean ± SD, n = 5, mm)

[0236]

[0237] Note: #: Compared with the normal control group, ##p<0.01; *: Compared with the model group, **p<0.01, ***p<0.001.

[0238] Mouse hair weight

[0239] At day 20, the weight of newly grown hair in the model group was significantly lower than that in the normal control group (p < 0.01). Compared with the model group, the weight of newly grown hair in the positive drug (minoxidil) group and the test drug (WH1) group was significantly increased (p < 0.01). It is worth noting that the weight of newly grown hair in the test drug (WH1) group was greater than that in the positive drug (minoxidil) group, but there was no statistical difference. This is likely because the number of mice in each group was small. The results are shown in Table 15.

[0240] Table 15 Hair weight of mice in each group after drug administration (mean ± SD, n = 5, mg)

[0241]

[0242] Note: #: Compared with the normal control group, ##p<0.01; *: Compared with the model group, **p<0.01, ***p<0.001.

[0243] HE staining results

[0244] In the normal group of mice, the skin tissue structure was intact and clearly layered, with the stratified keratinized squamous epithelium of the epidermis arranged neatly, without any atypia or abnormal proliferation. Accessory structures such as hair follicles, sebaceous glands, and eccrine sweat glands were visible in the dermis, with normal blood vessel distribution and no inflammatory cell infiltration or hemorrhage. In the model group of mice, the number of hair follicles in the dermis was significantly reduced, and the hair follicle structure was significantly atrophied. The large, deep-rooted terminal hair follicles were transformed into small, superficial vellus hair follicles. The lower segment of the hair follicle was shortened, the hair bulb was underdeveloped, the dermal papilla structure was blurred, and mild lymphocyte infiltration was visible around some hair follicles. The sebaceous glands were slightly larger than normal. In the positive control group, the epidermis of mice was clearly defined and of moderate thickness, with no abnormal keratinization or atrophy. The number of hair follicles in the dermis was significantly increased compared to the model group, with larger follicles, regular morphology, and prominent dermal papillae. The hair bulbs penetrated deep into the dermis or subcutaneous tissue, and the outer sheath cells of some hair follicles were densely arranged. No obvious inflammatory cell infiltration or tissue destruction was observed around the hair follicles, and the number of sebaceous glands was reduced compared to the model group. In the test drug group, the skin tissue structure of mice was improved compared to the model group. The number of hair follicles in the dermis was slightly increased compared to the model group, some hair follicles were larger, the lower segment of the hair follicles was extended and deepened, some hair follicles were irregular in shape, the hair bulbs were smaller, the dermal papillae were not clearly defined, and the inflammatory cell infiltration around the hair follicles was reduced compared to the model group. The results are shown in the figure. Figure 6 .

[0245] Compared with the normal group, the number of hair follicles, hair follicle length, hair bulb diameter, dermal thickness, and number of hair follicles in the anagen phase were significantly decreased in the model group (p < 0.01), while the number of hair follicles in the telogen phase was significantly increased (p < 0.01), indicating successful model establishment. Compared with the model group, the number of hair follicles, hair follicle length, hair bulb diameter, dermal thickness, and number of hair follicles in the anagen phase were significantly increased in the positive control group (minoxidil) and the test drug group (WH1) (p < 0.01), while the number of hair follicles in the telogen phase was significantly decreased (p < 0.01). The results are shown in Table 16. Figure 6 .

[0246] Table 16. Number of hair follicles, hair follicle length, hair bulb diameter, dermal thickness, number of hair follicles in the anagen phase, and number of hair follicles in the telogen phase in mice of each group after drug administration (mean ± SD, N = 3, 200 × 10⁻⁶).

[0247]

[0248] Research Results

[0249] During the administration period, at D19, one mouse in the model group (G2) died. The body weight of the dead mouse did not decrease significantly, and no abnormalities were found in the major organs upon gross dissection. The cause is considered to be the continuous administration of androgens that caused the death of the mouse. The remaining mice in the group did not decrease significantly in body weight and no other abnormalities were observed. The mice in the other groups were in good general condition, with no significant decrease in body weight and no other abnormalities were observed.

[0250] At days 14 and 20, the length of newly grown hair in the model group was significantly shorter than that in the normal control group (p < 0.01), indicating that the androgenic alopecia model was successful. At days 14 and 20, compared with the model group, the length of newly grown hair in the positive control group (minoxidil) and the test drug group (WH1) was significantly increased (p < 0.01). It is worth noting that at days 14 and 20, the length of newly grown hair in the test drug group (WH1) was greater than that in the positive control group (minoxidil), but there was no statistical difference, which is considered to be due to the small number of mice in each group.

[0251] At day 20, the weight of newly grown hair in the model group was significantly lower than that in the normal control group (p < 0.01). Compared with the model group, the weight of newly grown hair in the positive drug (minoxidil) group and the test drug (WH1) group was significantly increased (p < 0.01). It is worth noting that the weight of newly grown hair in the test drug (WH1) group was greater than that in the positive drug (minoxidil) group, but there was no statistical difference. This is likely due to the smaller number of mice in each group.

[0252] In the normal group of mice, the skin tissue structure was intact and clearly layered, with the stratified keratinized squamous epithelium of the epidermis arranged neatly, without any atypia or abnormal proliferation. Accessory structures such as hair follicles, sebaceous glands, and eccrine sweat glands were visible in the dermis, with normal blood vessel distribution and no inflammatory cell infiltration or hemorrhage. In the model group of mice, the number of hair follicles in the dermis was significantly reduced, and the hair follicle structure was significantly atrophied. The large, deep-rooted terminal hair follicles were transformed into small, superficial vellus hair follicles. The lower segment of the hair follicle was shortened, the hair bulb was underdeveloped, the dermal papilla structure was blurred, and mild lymphocyte infiltration was visible around some hair follicles. The sebaceous glands were slightly larger than normal. In the positive drug group, the epidermis of mice was clearly defined and of moderate thickness, with no abnormal keratinization or atrophy. The number of hair follicles in the dermis was significantly increased compared to the model group, with larger follicles, regular morphology, and prominent dermal papillae. The hair bulbs extended deep into the dermis or subcutaneous tissue, and the outer sheath cells of some hair follicles were densely arranged. No obvious inflammatory cell infiltration or tissue destruction was observed around the hair follicles, and the number of sebaceous glands was reduced compared to the model group. In the test drug group, the skin tissue structure of mice was improved compared to the model group. The number of hair follicles in the dermis was slightly increased compared to the model group, some hair follicles were larger, the lower segment of the hair follicles was extended and deepened, some hair follicles were irregular in shape, the hair bulbs were smaller, the dermal papillae were not clear, and the inflammatory cell infiltration around the hair follicles was reduced compared to the model group.

[0253] Compared with the normal group, the number of hair follicles, hair follicle length, hair bulb diameter, dermal thickness, and number of hair follicles in the anagen phase were significantly reduced in the model group (p < 0.01), while the number of hair follicles in the telogen phase was significantly increased (p < 0.01), indicating that the model was successfully established. Compared with the model group, the number of hair follicles, hair follicle length, hair bulb diameter, dermal thickness, and number of hair follicles in the anagen phase were significantly reduced in the positive control group (minoxidil) and the test group (WH1) (p < 0.01), while the number of hair follicles in the telogen phase was significantly reduced (p < 0.01).

[0254] In a mouse model of androgenetic alopecia, the test drug WH1 significantly promoted hair growth, demonstrating a marked therapeutic effect on androgenetic alopecia. Furthermore, under the experimental conditions, mice tolerated WH1 well. The compound provided by this invention targets the SFRP1 protein in the Wnt / β-catenin pathway, exhibiting a strong affinity for SFRP1, thereby activating the Wnt / β-catenin signaling pathway, further promoting hair follicle cell growth and hair development. It also exhibits minimal skin irritation and good tolerability, achieving an effective treatment for androgenetic alopecia.

[0255] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A compound for use in the treatment of androgenetic alopecia, characterized in that, having a structure represented by any one of formula (I) - formula (IV): wherein R is one of methyl, alkoxy, trifluoromethyl, halogen, carbonamide, sulfonamide, amino, and sulfone; X and Y are independently selected from carbon atom or nitrogen atom; W is carbon atom or nitrogen atom.

2. Use of a compound for treating androgenetic alopecia as a specific inhibitor of SFRP1 according to claim 1.

3. Use of a compound for treating androgenetic alopecia in the manufacture of a medicament for treating androgenetic alopecia according to claim 1.

4. A pharmaceutical composition for treating androgenetic alopecia, characterized by, Compounds comprising a structure represented by any one of formula (I) - formula (IV) in claim 1 as a pharmaceutical active ingredient.

5. The pharmaceutical composition of claim 4, wherein, The dosage form of the pharmaceutical composition includes aerosol, solution, gel, cream, ointment, or film.

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