A pharmaceutical composition containing exosomes and uses thereof

Abstract

The application belongs to the technical field of biotechnology, and particularly relates to a pharmaceutical composition containing exosomes and application thereof. The pharmaceutical composition containing exosomes comprises stem cell exosomes, recombinant IGFBP7 protein and baeckea fruitin B; the amino acid sequence of the recombinant IGFBP7 protein is shown as SEQ ID NO:1. Test results show that the combination of umbilical cord mesenchymal stem cell exosomes, baeckea fruitin B and recombinant IGFBP7 protein can stimulate hair follicle stem cells, prolong the initial stage of hair growth of hair follicles, and thus effectively improve the hair loss of patients.

Description

Technical Field

[0001] This invention belongs to the field of biotechnology, specifically relating to a pharmaceutical composition containing exosomes and its application. Background Technology

[0002] Currently, there are two main treatment methods for hair loss: medication and hair transplantation. Rogaine, a hair loss drug approved by the U.S. Food and Drug Administration (FDA), is the only proven effective hair regrowth drug on the market. However, its effects are limited to the short term and cannot fundamentally improve the scalp's microenvironment. Once discontinued, hair growth quickly reverts to its pre-treatment state, and long-term use may lead to side effects such as hormonal imbalances. On the other hand, hair transplantation, as an invasive and expensive treatment, while feasible, does not alter the microenvironment of the hair follicles. Therefore, newly transplanted hair is prone to falling out again, failing to address the root cause of the problem. It is worth noting that stem cell exosome products are currently mostly used in tissue repair and cosmetic fields, and their application in hair loss prevention and treatment, especially in the treatment of common androgenetic alopecia, is still rare.

[0003] Chronic inflammation around hair follicles is known to be a significant factor contributing to androgenetic alopecia. Inflammation leads to apoptosis of hair follicle cells, further accelerating follicle degeneration and atrophy, thus affecting hair growth. Anti-inflammatory treatment may be an effective approach for addressing the inflammatory factors in androgenetic alopecia. By using anti-inflammatory drugs or physical therapy, the inflammatory response around hair follicles can be reduced, thereby slowing the progression of hair loss. Traditional Chinese medicine extracts have high activity and low toxicity, showing potential for treating hair loss. While gentian root extract (e.g., gentian root extract) exhibits strong anti-inflammatory activity, research on its use in treating androgenetic alopecia remains incomplete.

[0004] Furthermore, studies have reported lower levels of IGFBP7 in the serum and scalp hair follicles of AGA patients compared to healthy controls. Subcutaneous injection of IGFBP7 delayed the onset of regression in DHT-induced AGA mice and prolonged the early stage of hair growth in scalp hair follicles, demonstrating significant therapeutic potential for AGA. However, existing recombinant IGFBP7 proteins have very complex structures, making artificial expression and large-scale production difficult and costly, and their stability is poor. Therefore, developing recombinant IGFBP7 proteins with smaller molecular weights and higher stability has become a crucial issue that urgently needs to be addressed. Summary of the Invention

[0005] The primary objective of this invention is to provide a pharmaceutical composition containing exosomes that can stimulate hair follicle stem cells, prolong the early stage of hair growth in hair follicles, and thus effectively improve hair loss in AGA patients.

[0006] The second objective of this invention is to provide an application of a pharmaceutical composition containing exosomes, which has broad application prospects.

[0007] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0008] A pharmaceutical composition containing exosomes, the pharmaceutical composition comprising stem cell exosomes, recombinant IGFBP7 protein and sucralose; the amino acid sequence of the recombinant IGFBP7 protein is shown in SEQ ID NO:1.

[0009] The sequence of SEQ ID NO:1 is as follows:

[0010] PGVSGVCVCKSGYPVCGSDGLAYPSGCQLRAASQRAESRGEGAITQVHKGTCEQPHIV

[0011] TPPKDIWNVTGAQVYLSCEVCGIPTPVLIWNKVKRGHYGVQRTELLPGDRDNLHGQTRGG

[0012] PEKHEVTGWVLVSPLSKEDAGEYECHASNSAGQASASAKHT

[0013] Furthermore, the nucleotide sequence of the recombinant IGFBP7 protein is shown in SEQ ID NO:2.

[0014] The sequence of SEQ ID NO:2 is as follows:

[0015] CCGGGCGTATCTGGTGTATGTGTCTGCAAGTCTGGCTACCCGGTGTGCGGCAGCGA

[0016] CGGCCTGGCCTACCCCAGCGGCTGCCAGCTGAGAGCCGCCAGCCAGAGAGCCGAGAGC

[0017] AGAGGCGAGGGCGCCATCACCCAGGTGCACAAGGGCACCTGCGAGCAGCCCCACATCG

[0018] TGACCCCCCCCCAAGGACATCTGGAACGTGACCGGCGCCCAGGTGTACCTGAGCTGCGA

[0019] GGTGTGCGGCATCCCCACCCCCGTGCTGATCTGGAACAAGGTGAAGAGAGGCCACTAC

[0020] GGCGTGCAGAGAACCGAGCTGCTGCCCGGCGACAGAGACAACCTGCACGGCCAGACC

[0021] AGAGGCGGCCCCGAGAAGCACGAGGTGACCGGCTGGGTGCTGGTGAGCCCCTGAGC

[0022] AAGGAGGACGCCGGCGAGTACGAGTGCCACGCCAGCAACAGCGCCGGCCAGGCCAGC

[0023] GCCAGCGCCAAGCACACC

[0024] Furthermore, the mass ratio of stem cell exosomes, recombinant IGFBP7 protein, and sucralose in the pharmaceutical composition is 1:(1-2):1.

[0025] Furthermore, the method for preparing the recombinant IGFBP7 protein is as follows:

[0026] (1) The nucleotide sequence encoding the recombinant IGFBP7 protein is ligated to a vector to obtain a recombinant plasmid;

[0027] (2) Transform the recombinant plasmid from step (1) into bacteria to construct recombinant bacteria;

[0028] (3) The recombinant bacteria constructed in step (2) were expressed. The bacterial precipitate was collected, the supernatant was obtained by cleavage, and the recombinant IGFBP7 protein was obtained by purification.

[0029] Further, the expression process in step (3) is as follows: the recombinant bacteria are inoculated into LB medium containing kanamycin and shaken overnight, and IPTG is added to induce the expression of recombinant IGFBP7 protein; the induction concentration of IPTG is 0.3-0.5 mmol / L, the induction time of IPTG is 5-10 h, and the concentration of kanamycin is 60-90 μg / mL.

[0030] Furthermore, the stem cell exosomes are umbilical cord mesenchymal stem cell exosomes.

[0031] Furthermore, the method for obtaining the umbilical cord mesenchymal stem cell exosomes is as follows:

[0032] According to (1-5)×10 4 Umbilical cord mesenchymal stem cells (UMCs) were seeded into culture medium at a density of P2-P5 generation cells / mL and cultured. The cell culture supernatant was collected, and the cell culture supernatant was separated by ultracentrifugation. The precipitate was collected and freeze-dried to obtain UMC exosomes.

[0033] Furthermore, the culture medium consists of DMEM / F12 basal medium and the following components added to the medium: 8-10% FBS, 90-100 ng / mL IL-2, 30-50 ng / mL IL-21, and 1-5 ng / mL EGF; the culture time is 2-3 days, and the culture conditions are 37°C and 5% CO2.

[0034] The application of the above-mentioned pharmaceutical composition containing exosomes in the preparation of products for treating hair loss.

[0035] Furthermore, the amount of the pharmaceutical composition added is 2-8% based on the total mass of the product.

[0036] Compared with the prior art, the main advantages of the present invention are as follows:

[0037] This invention provides a pharmaceutical composition comprising umbilical cord mesenchymal stem cell exosomes, recombinant IGFBP7 protein, and ethyl chernozepine. The recombinant IGFBP7 protein is obtained by effectively truncating and designing the amino acid sequence of the full-length human IGFBP7 protein. This recombinant IGFBP7 protein has a smaller molecular weight, better stability, and stronger activity. Experimental results show that the combined use of umbilical cord mesenchymal stem cell exosomes, ethyl chernozepine, and recombinant IGFBP7 protein can stimulate hair follicle stem cells, prolonging the early stage of hair growth in hair follicles, thereby effectively improving hair loss in AGA patients. Specifically, umbilical cord mesenchymal stem cell exosomes can promote hair follicle repair and regeneration, shortening the telogen phase of hair follicles; ethyl chernozepine can reduce inflammatory responses and improve local microcirculation, thereby accelerating the entry of telogen follicles into the anagen phase. The recombinant IGFBP7 protein of this invention can better promote the in vitro migration of human outer root sheath cells and prolong the early stage of hair growth in hair follicles. Attached Figure Description

[0038] Figure 1 This is a morphological diagram of P3 generation umbilical cord mesenchymal stem cells;

[0039] Figure 2 This is a graph showing the results of SDS-PAGE electrophoresis identification of recombinant IGFBP7 protein. Figure 2 In this context, "M" represents the protein marker, and "1" represents lane 1, which is the purified recombinant protein IGFBP7. Detailed Implementation

[0040] The technical solution of the present invention will be further described below with reference to specific embodiments. However, those skilled in the art should understand that the following embodiments are only for illustrating the present invention and should not be regarded as limiting the present invention. Specific conditions not specified in the embodiments are performed according to conventional conditions or conditions recommended by the manufacturer. Unless otherwise specified, the reagents or instruments used are all conventional products obtained through commercial channels.

[0041] The catalog number of the commercially available recombinant IGFBP7 protein of this invention is abs04667.

[0042] Example

[0043] Example 1

[0044] Example 1: Obtaining exosomes from umbilical cord mesenchymal stem cells. The specific method is as follows:

[0045] Wash the umbilical cord tissue with PBS buffer and cut it into 1mm pieces with surgical scissors. 3 The tissue samples were plated in 10cm culture dishes (serum-free DMEM / F12 medium containing 10% fetal bovine serum) and cultured at 37℃ in a 5% CO2 cell culture incubator until the cells reached 80% confluence. Primary cultured umbilical cord mesenchymal stem cells were obtained by digestion with Tryspin-EDTA enzyme. The cells were then cultured at a rate of 3 × 10⁻⁶ cells / year. 4 Primary cultured umbilical cord mesenchymal stem cells were seeded into a culture medium consisting of DMEM / F12 basal medium and the following components: 10% FBS, 100 ng / mL IL-2, 50 ng / mL IL-21, and 5 ng / mL EGF. When the cell confluence reached 90%, the cells were digested with trypsin solution containing 0.25% EDTA and passaged in a 37°C, 5% CO2 cell culture incubator at a passage ratio of 1:2 for 3 consecutive passages to obtain P3 generation umbilical cord mesenchymal stem cells. The morphology of P3 generation umbilical cord mesenchymal stem cells is shown in the figure below. Figure 1 As shown.

[0046] According to 3×10 4P3 generation umbilical cord mesenchymal stem cells were seeded into a culture medium consisting of DMEM / F12 basal medium and the following components added to the medium: 10% FBS, 100 ng / mL IL-2, 50 ng / mL IL-21, and 5 ng / mL EGF. The cells were cultured at 37°C in a 5% CO2 cell culture incubator for 2 days, and the cell culture supernatant was collected. The cell supernatant was centrifuged at 400g for 8 min, then centrifuged at 4500g for 6 min, and then centrifuged at 85000g for 30 min. The precipitate was collected, resuspended, and then centrifuged at 85000g for 30 min. After freeze-drying, umbilical cord mesenchymal stem cell exosomes were obtained.

[0047] Example 2

[0048] Example 2: Obtaining recombinant IGFBP7 protein. The specific preparation process is as follows:

[0049] (1) Based on the amino acid sequence of the full-length human IGFBP7 protein, functional fragments were cut and designed to obtain the amino acid sequence of the recombinant IGFBP7 protein as shown in SEQ ID NO:1; the gene sequence was reverse-engineered based on the amino acid sequence of the recombinant IGFBP7 protein, and the corresponding base sequence was codon optimized according to the codon preference of E. coli in expressing heterologous proteins to obtain the nucleotide sequence of the recombinant IGFBP7 protein as shown in SEQ ID NO:2; the nucleotide sequence of the recombinant IGFBP7 protein was ligated with the pET28a vector, and the recombinant plasmid IGFBP7-pET28a was obtained after identification and sequencing;

[0050] The sequence of SEQ ID NO:1 is as follows:

[0051] PGVSGVCVCKSGYPVCGSDGLAYPSGCQLRAASQRAESRGEGAITQVHKGTCEQPHIV

[0052] TPPKDIWNVTGAQVYLSCEVCGIPTPVLIWNKVKRGHYGVQRTELLPGDRDNLHGQTRGG

[0053] PEKHEVTGWVLVSPLSKEDAGEYECHASNSAGQASASAKHT

[0054] The sequence of SEQ ID NO:2 is as follows:

[0055] CCGGGCGTATCTGGTGTATGTGTCTGCAAGTCTGGCTACCCGGTGTGCGGCAGCGACGGCCTGGCCTACCCCAGCGGCTGCCAGCTGAGAGCCGCCAGCCAGAGAGCCGAGAGCAGAGGCGAGGGCGCCATCACCCAGGTGCACAAGGGCACCTGCGAGCAGCCCCACATCGTGACCCCCCAAGGACATCTGGAACGTGACCGGCGCCCAGGTGTACCTGAGCTGCGAGGTGTGCG GCATCCCCACCCCCGTGCTGATCTGGAACAAGGTGAAGAGAGGCCACTACGGCGTGCAGAGAACCGAGCTGCTGCCCGGCGACAGAGACAACCTGCACGGCCAGACCAGAGGCGGCCCCGAGAAGCACGAGGTGACCGGCTGGGTGCTGGTGAGCCCCCTGAGCAAGGAGGACGCCGGCGAGTACGAGTGCCACGCCAGCAACAGCGCCGGCCAGGCCAGCGCCAGCGCCAAGCACACC

[0056] (2) The recombinant plasmid obtained in step (1) was transformed into E. coli BL21(DE3) engineered bacteria. The recombinant bacteria were evenly spread on LB agar plates containing 70 μg / mL kanamycin and incubated overnight at 37°C to obtain recombinant bacteria IGFBP7-pET28a / BL21.

[0057] (3) Select a single positive colony of the recombinant strain IGFBP7-pET28a / BL21 from step (2) and inoculate it into 4 mL of LB medium containing 50 μg / mL kanamycin. Incubate overnight at 37°C with shaking at 230 rpm. Then, inoculate 1% of the total culture volume of the colony into LB medium containing 70 μg / mL kanamycin and incubate at 37°C with shaking at 220 rpm for 3 h until OD500 reaches 100%. 600 The concentration of IPTG was 0.6, and a final concentration of 0.4 mM was added. The mixture was induced at 16℃ and 180 rpm for 8 h. The bacterial cells were then collected by centrifugation. Crude protein was obtained by separation and affinity chromatography, followed by ion exchange chromatography to obtain purified recombinant IGFBP7 protein. The purity of the recombinant protein was identified by SDS-PAGE electrophoresis, and the results are as follows: Figure 2 As shown.

[0058] Figure 2 This is a graph showing the results of SDS-PAGE electrophoresis identification of recombinant IGFBP7 protein. From... Figure 2It can be seen that the recombinant IGFBP7 protein has high purity, the estimated molecular weight of the recombinant protein is 17kDa, and the band position is consistent with the expected size.

[0059] Example 3

[0060] Example 3 provides a pharmaceutical composition containing exosomes, which consists of umbilical cord mesenchymal stem cell exosomes prepared in Example 1, recombinant IGFBP7 protein prepared in Example 2, and sucralose ethyl, wherein the mass ratio of umbilical cord mesenchymal stem cell exosomes, recombinant IGFBP7 protein, and sucralose ethyl is 1:1:1.

[0061] Example 4

[0062] Example 4 provides a pharmaceutical composition containing exosomes, which consists of umbilical cord mesenchymal stem cell exosomes prepared in Example 1, recombinant IGFBP7 protein prepared in Example 2, and sucralose, wherein the mass ratio of umbilical cord mesenchymal stem cell exosomes, recombinant IGFBP7 protein, and sucralose is 1:1.5:1.

[0063] Example 5

[0064] Example 5 provides a pharmaceutical composition containing exosomes, which consists of umbilical cord mesenchymal stem cell exosomes obtained in Example 1, recombinant IGFBP7 protein obtained in Example 2, and bilirubin B, wherein the mass ratio of umbilical cord mesenchymal stem cell exosomes, recombinant IGFBP7 protein, and bilirubin B is 1:2:1.

[0065] Comparative Example

[0066] Comparative Example 1

[0067] This comparative example provides a pharmaceutical composition that differs from Example 4 in that: the pharmaceutical composition of this comparative example consists of umbilical cord mesenchymal stem cell exosomes prepared in Example 1 and recombinant IGFBP7 protein prepared in Example 2, wherein the mass ratio of umbilical cord mesenchymal stem cell exosomes to recombinant IGFBP7 protein is 1:1.5.

[0068] Comparative Example 2

[0069] This comparative example provides a drug that differs from Example 4 in that the drug in this comparative example is the umbilical cord mesenchymal stem cell exosome prepared in Example 1, and the amount of umbilical cord mesenchymal stem cell exosome is the same as the total amount of umbilical cord mesenchymal stem cell exosome, recombinant IGFBP7 protein, and sucralose in Example 4.

[0070] Comparative Example 3

[0071] This comparative example provides a drug that differs from Example 4 in that the drug in this comparative example is the recombinant IGFBP7 protein prepared in Example 2, and the amount of recombinant IGFBP7 protein used is the same as the total amount of umbilical cord mesenchymal stem cell exosomes, recombinant IGFBP7 protein, and sucralose in Example 4.

[0072] Comparative Example 4

[0073] This comparative example provides a pharmaceutical composition that differs from Example 4 in that: the pharmaceutical composition of this comparative example consists of umbilical cord mesenchymal stem cell exosomes obtained in Example 1, commercially available recombinant IGFBP7 protein, and bilirubin B, wherein the mass ratio of umbilical cord mesenchymal stem cell exosomes, commercially available recombinant IGFBP7 protein, and bilirubin B is 1:1.5:1.

[0074] The properties of the pharmaceutical compositions of the present invention are described below.

[0075] The effects of the drugs obtained in Examples 3-5 and Comparative Examples 1-4 were tested, as follows:

[0076] Experimental subjects: 100 male SPF-grade C57BL / 6 mice aged 6 weeks.

[0077] Rearing environment: temperature (23±2)℃, relative humidity 40%-60%, light and darkness 12h each.

[0078] AGA model establishment: All mice were acclimatized for one week and anesthetized with ether. Rosin and paraffin were heated and melted (1:1), cooled, and applied to a 2cm × 2cm area on the back of the mice. After solidification, the mixture was removed to confirm that the mouse hair was in the resting phase (skin turned pink). On the second day, the mice were randomly divided into a blank group, a model group, Examples 3-5, and Comparative Examples 1-4, with 10 mice in each group.

[0079] Administration: Except for the control group, 200 μL of 0.05% testosterone (prepared with 75% ethanol) was applied to the hair removal area of ​​the other groups once a day for 28 consecutive days. After 30 min of testosterone treatment, 200 μL of the drug was administered in the same manner. The control group and the model group were given 75% ethanol, while the other groups were given 40 mg / mL of the drug used in Examples 3-5 and Comparative Examples 1-4 (all prepared with 75% ethanol).

[0080] Before applying the medication each day, observe the skin on the backs of mice in each group and record the time when the skin discoloration (beginning to turn gray) is recorded. After 28 days of treatment, euthanize the mice by cervical dislocation, and pluck hair from five areas of the bald zone (upper left, lower left, middle, upper right, and lower right) and measure the weight using calipers. Also, shave the hair from the bald zones and weigh the hair. The final data is the average of the test data for each group, and the results are shown in Table 1.

[0081] Table 1

[0082]

[0083]

[0084] As can be seen from the above, compared with the blank group, the skin discoloration time and hair growth time of the model group mice were significantly delayed, and the hair length was shorter and the hair quality was lighter.

[0085] Compared to the model group, after using the drugs obtained in Examples 3-5, the mice showed a 2-day reduction in skin discoloration time and hair growth time, and a significant increase in hair length and quality. These results indicate that the drugs obtained in Examples 3-5 can alleviate the inhibitory effect of testosterone on hair growth in C57BL / 6 mice.

[0086] The effects of the drugs obtained in Comparative Examples 1-3 were not as good as those in Examples 3-5. Analysis revealed that Comparative Example 1 did not contain cypermethrin; Comparative Example 2 did not contain recombinant IGFBP7 protein or cypermethrin, but increased the amount of umbilical cord mesenchymal stem cell exosomes; Comparative Example 3 did not contain umbilical cord mesenchymal stem cell exosomes or cypermethrin, but increased the amount of recombinant IGFBP7 protein. Therefore, the above results indicate that the combined use of umbilical cord mesenchymal stem cell exosomes, cypermethrin, and recombinant IGFBP7 protein can stimulate hair follicle stem cells, prolonging the early stage of hair growth in hair follicles, thereby effectively improving hair loss in AGA patients. The reason for this may be that umbilical cord mesenchymal stem cell exosomes can promote hair follicle repair and regeneration, shortening the telogen phase; cypermethrin can reduce inflammatory response and improve local microcirculation, thereby accelerating the entry of telogen phase hair follicles into the anagen phase. The recombinant IGFBP7 protein of this invention can better promote the in vitro migration of human outer root sheath cells, prolonging the early stage of hair growth in hair follicles.

[0087] Compared to Examples 3-5, the skin and hair growth on the backs of mice were slightly worse after using the drug obtained in Comparative Example 4. This may be because Comparative Example 4 used commercially available recombinant IGFBP7 protein instead of the recombinant IGFBP7 protein prepared in this invention. Compared to commercially available recombinant IGFBP7 protein, the recombinant IGFBP7 protein obtained in this invention has a smaller molecular weight, better activity and stability, and can thus more effectively alleviate the inhibitory effect of testosterone on hair growth in C57BL / 6 mice.

[0088] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. The basic principles and main features of the present invention have been described above with specific implementation schemes. Based on the present invention, some modifications or substitutions can be made, but these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of protection claimed by the present invention.

Claims

1. A pharmaceutical composition containing exosomes, characterized in that, The pharmaceutical composition comprises stem cell exosomes, recombinant IGFBP7 protein, and sucralose; the amino acid sequence of the recombinant IGFBP7 protein is shown in SEQ ID NO:

1. The mass ratio of stem cell exosomes, recombinant IGFBP7 protein, and ethyl succinate in the pharmaceutical composition is 1:(1-2):1; The method for preparing the recombinant IGFBP7 protein is as follows: (1) The nucleotide sequence encoding the recombinant IGFBP7 protein is ligated to a vector to obtain a recombinant plasmid; (2) Transform the recombinant plasmid from step (1) into bacteria to construct recombinant bacteria; (3) The recombinant bacteria constructed in step (2) were expressed, the bacterial precipitate was collected, the supernatant was obtained by cleavage, and the recombinant IGFBP7 protein was obtained by purification. The expression process described in step (3) is as follows: the recombinant bacteria are inoculated into LB medium containing kanamycin and shaken overnight, and IPTG is added to induce the expression of recombinant IGFBP7 protein; the IPTG induction concentration is 0.3-0.5 mmol / L and the IPTG induction time is 5-10 h; The stem cell exosomes are umbilical cord mesenchymal stem cell exosomes; The method for obtaining the umbilical cord mesenchymal stem cell exosomes is as follows: According to (1-5)×10 4 P2-P5 generation umbilical cord mesenchymal stem cells were seeded into culture medium at a density of cells / mL and cultured. The cell culture supernatant was collected, and the cell culture supernatant was separated by ultracentrifugation. The precipitate was collected and freeze-dried to obtain umbilical cord mesenchymal stem cell exosomes. The culture medium consists of DMEM / F12 basal medium and the following components added to the medium: 8-10% FBS, 90-100 ng / mL IL-2, 30-50 ng / mL IL-21, and 1-5 ng / mL EGF; the culture time is 2-3 days, and the culture conditions are 37℃ and 5% CO2.

2. The pharmaceutical composition containing exosomes according to claim 1, characterized in that, The nucleotide sequence of the recombinant IGFBP7 protein is shown in SEQ ID NO:

2.

3. The pharmaceutical composition containing exosomes according to claim 1, characterized in that, The concentration of kanamycin in step (3) is 60-90 μg / mL.

4. The use of the pharmaceutical composition containing exosomes according to any one of claims 1-3 in the preparation of a product for treating androgenetic alopecia.

5. The use of the pharmaceutical composition containing exosomes according to claim 4 in the preparation of a product for treating androgenetic alopecia, characterized in that, The amount of the pharmaceutical composition added is 2-8% based on the total mass of the product.

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