Application of small molecular compound and analogue thereof in treatment of ovarian senescence diseases

Abstract

The invention discloses an application of a small molecular compound and an analogue thereof in treating ovarian aging diseases. It is found for the first time that the exogenous supplementary adenosine or the analogues thereof can effectively recover or improve the ovarian function and can prevent and inhibit ovarian fibrosis, and therefore the exogenous supplementary adenosine or the analogues thereof can be used as potential drugs for recovering the female fertility.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and in particular to the application of a small molecule compound and its analogues in the treatment of ovarian aging diseases. Background Technology

[0002] The ovary is the primary reproductive organ of female mammals, responsible for producing eggs and secreting sex hormones. Normal ovarian function directly affects the fertility and reproductive health of female animals. Aging is one of the main causes of ovarian dysfunction, but environmental factors, genetic factors, and diseases can also affect ovarian function, leading to a gradual increase in the incidence of ovarian dysfunction and potentially causing a series of reproductive system-related diseases.

[0003] Adenosine (ade) is an endogenous nucleoside widely distributed in organisms, possessing various physiological functions including regulating cellular metabolism, immune responses, and neurotransmission. Studies have shown that adenosine and its receptors also play an important role in the reproductive system. However, there are currently no clear reports on the application of adenosine in restoring ovarian function.

[0004] Therefore, in order to solve the above problems, developing a drug-based treatment for infertility caused by ovarian dysfunction is of great significance to clinical medicine and infertile patients. Summary of the Invention

[0005] This invention aims to at least solve one of the aforementioned technical problems existing in the prior art. Therefore, the object of this invention is to provide the application of small molecule compounds and their analogues in the treatment of ovarian aging diseases. This invention is the first to discover that exogenous supplementation of adenosine or its analogues can effectively restore or improve ovarian function and can prevent and inhibit ovarian fibrosis, thus it can be used as a potential drug for restoring female fertility.

[0006] In a first aspect, the invention provides the use of compounds of Formula I in the preparation of medicaments for the prevention and / or treatment of ovarian fibrosis.

[0007]

[0008] Formula I.

[0009] In some embodiments of the present invention, R is selected from O or CH2.

[0010] In this invention, the term "ovarian fibrosis" refers to age-related ovarian fibrosis characterized by higher levels of permanent collagen deposition, which leads to impaired ovulation and damages ovarian function.

[0011] In some embodiments of the present invention, the compound is adenosine or a pharmaceutically acceptable salt thereof, or mongolicin or a pharmaceutically acceptable salt thereof.

[0012] In some embodiments of the present invention, the pharmaceutically acceptable salts include, but are not limited to: anionic salts, such as hydrochloride, acetate, fumarate, maleate, citrate, etc.; and cationic salts, such as sodium salts, potassium salts, meglumine salts, etc.

[0013] In some embodiments of the present invention, the medicament further contains pharmaceutically acceptable excipients.

[0014] In some embodiments of the present invention, the excipients include at least one of diluents, absorbents, binders, disintegrants, lubricants, colorants, coating materials, solvents, pH adjusters, antibacterial agents, isotonic adjusters, and chelating agents.

[0015] In some embodiments of the present invention, the diluent includes, but is not limited to, starch, dextrin, sucrose, lactose, mannitol, etc.

[0016] In some embodiments of the present invention, the absorbent includes, but is not limited to, calcium sulfate, calcium hydrogen phosphate, etc.

[0017] In some embodiments of the present invention, the wetting agent includes, but is not limited to, ethanol.

[0018] In some embodiments of the present invention, the adhesive includes, but is not limited to, hydroxypropyl methylcellulose, povidone, etc.

[0019] In some embodiments of the present invention, the disintegrant includes, but is not limited to, sodium hydroxymethyl starch, crospovidone, etc.

[0020] In some embodiments of the present invention, the lubricant includes, but is not limited to, talc, hydrogenated vegetable oil, polyethylene glycol, etc.

[0021] In some embodiments of the present invention, the colorant includes, but is not limited to, titanium dioxide, methylene blue, etc.

[0022] In some embodiments of the present invention, the antibacterial agent includes, but is not limited to, sodium sulfite, sodium thiosulfate, etc.

[0023] In some embodiments of the present invention, the isotonic regulator includes, but is not limited to, glucose, sodium chloride, etc.

[0024] In some embodiments of the present invention, the chelating agent includes, but is not limited to, disodium EDTA.

[0025] In some embodiments of the present invention, the drug further contains a second active substance.

[0026] In some embodiments of the present invention, the second active substance has at least one of the following functions (1)-(5): (1) Improve or restore ovarian function; (2) To treat or improve menstrual disorders; (3) Increase or improve estrogen (E2) levels; (4) Increase or improve anti-Müllerian hormone (AMH) levels; (5) Improve fertilization rate, high-quality embryo rate or egg retrieval rate.

[0027] A second aspect of the present invention provides the use of compounds with the structural formula shown in Formula I in the preparation of aphrodisiac drugs;

[0028] Formula I; R is selected from O or CH2.

[0029] In some embodiments of the present invention, the compounds are defined as described above.

[0030] In some embodiments of the present invention, the drug is defined as described above.

[0031] In this invention, the term "aphrodisiac" refers to enhancing the estrous cycle of the subject.

[0032] In some embodiments of the present invention, the subjects include humans (females) and non-human female mammals.

[0033] In this invention, mice, which share common characteristics with humans, are used as experimental subjects. Similar to the human menstrual cycle, mice also have regular estrous cycles accompanied by ovulation. The mouse estrous cycle consists of four phases: proestrus, estrus, metroestrus, and mitrus, with a normal estrous cycle lasting 3-5 days. Variations in the mouse estrous cycle are closely related to hormone secretion and ovarian function. Therefore, it can also reflect changes in ovarian function.

[0034] This invention uses "older" mice to simulate the decline in ovarian function caused by natural aging.

[0035] A third aspect of the invention provides the use of compounds of Formula I in the preparation of medicaments for enhancing or improving female reproductive capacity;

[0036] Formula I; R is selected from O or CH2.

[0037] In some embodiments of the present invention, the compounds are defined as described above.

[0038] In some embodiments of the present invention, the drug is defined as described above.

[0039] In some embodiments of the present invention, the improvement or enhancement of female reproductive capacity includes at least one of: enhancing ovarian reserve and increasing ovarian area.

[0040] In this invention, the term "ovarian reserve" refers to the primordial follicles contained in the ovarian cortex of human women. It also refers to the primordial follicles in the ovaries of mice. A follicle is the basic unit of fertility, consisting of an oocyte and surrounding layers of granulosa cells. Follicles can be classified into primordial follicles, primary follicles, secondary follicles, antral follicles, and atretic follicles according to their different developmental stages. The number of follicles at each stage and the total number of follicles can reflect fertility and ovarian function.

[0041] A fourth aspect of the present invention provides a method for improving fertility, comprising: Administer an effective amount of the compound with the structural formula shown in Formula I to the subject;

[0042] Formula I; R is selected from O or CH2.

[0043] In some embodiments of the present invention, the subject is a normal person or a patient with reproductive defects.

[0044] In some embodiments of the present invention, the method does not involve the treatment of a disease.

[0045] The beneficial effects of this invention are: This invention is the first to discover the effectiveness of adenosine or its analogues in restoring ovarian function. Through experimental verification, the adenosine and adenosine analogues of this invention have significant promoting effects on restoring ovarian function, restoring or improving the estrous cycle, and preventing ovarian fibrosis. Therefore, they can be used for ovarian disorders caused by chemotherapy or natural aging, providing a new drug option for relevant populations. Attached Figure Description

[0046] Figure 1 The results of follicle counts in eight-month-old mice one month after administration of adenosine.

[0047] Figure 2 The results of the mouse estrous cycle statistics are presented in Table 1.

[0048] Figure 3 The results show the ovarian area of ​​eight-month-old mice one month after administration of adenosine.

[0049] Figure 4The results show the ovarian fibrosis area of ​​mice treated with adenosine for 1-2 months. In A, the mice were 8 months old after 1 month of treatment, and in B, the mice were 12 months old after 2 months of treatment. Detailed Implementation

[0050] The present invention will be further described in detail below through specific embodiments. Unless otherwise specified, the raw materials, reagents, or apparatus used in the embodiments and comparative examples are all available from conventional commercial sources or can be obtained by existing technical methods. Unless otherwise specified, the test or experimental methods are conventional methods in the art.

[0051] Experimental materials The mice used in the following examples were 8-month-old and 12-month-old female C57BL / 6 mice, purchased from the Animal Center of Southern Medical University. All mice were housed in a specific pathogen-free environment at 22-25 degrees Celsius.

[0052] The adenosine and its analogues used were purchased from Sigma. The CAS number of adenosine was 58-61-7, and the CAS number of the adenosine analogue was 19186-33-5 (Mongrezin).

[0053] Example 1 This embodiment investigates the effect of adenosine on the recovery of ovarian function in aging mice. The specific experimental steps are as follows: Take 2 mg of adenosine powder and dissolve it completely in 1 mL of sterile saline (using ultrasound or at 37°C) to obtain a final concentration of 75 mM adenosine stock solution. Sterilize using a 0.22 μm pore size filter and dilute to the required concentration with sterile saline according to the application requirements.

[0054] Twenty-six 8-month-old and six 12-month-old female C57BL / 6 mice were randomly divided into a control group (Con) and a treatment group (Ade).

[0055] 10 μL of the adenosine stock solution was diluted in 9.99 mL of sterile saline, and the diluted solution was used for intraperitoneal injection (dose was 15 mg adenosine / kg mouse body weight), once daily. The mouse body weight was recorded after each administration.

[0056] During the last 10 days of drug administration, the estrous cycle and status of mice were monitored and recorded. The specific method was as follows: vaginal smears were prepared from samples taken from each mouse and stained with Gymza dye. Under a microscope, the estrous cycle was determined by cell type and proportion. Estrous cycle line graphs were plotted based on the records to determine if any abnormalities existed in the mouse's estrous cycle.

[0057] After the experimental period ended, 8-month-old and 12-month-old mice were subjected to the following treatments: For 8-month-old mice, after one month of continuous administration, the mice were anesthetized with 1% pentobarbital, and blood was collected by enucleation. The obtained blood samples were left at room temperature for 2 hours, and then centrifuged at 2000 g for 15 minutes at 4 degrees Celsius using a low-temperature centrifuge. The supernatant serum was collected for subsequent detection of serum hormone levels.

[0058] For 12-month-old mice, after two months of continuous administration, the mice were anesthetized with 1% pentobarbital, and blood was collected by enucleation. The obtained blood samples were left at room temperature for 2 hours, and then centrifuged at 2000 g for 15 minutes at 4 degrees Celsius using a low-temperature centrifuge. The supernatant serum was collected for subsequent detection of serum hormone levels.

[0059] Simultaneously, after euthanizing the mice by cervical dislocation, ovarian and liver tissues (including the lobe and gallbladder lobe) were collected from all mice. The tissues were fixed in five times their volume of 4% paraformaldehyde solution (12-24 h). After fixation, the tissues were embedded in paraffin and sectioned (5 μm thickness).

[0060] Tissue sections were stained with hematoxylin and eosin (HE) and phosphoric acid (PSR), and observed and photographed under a microscope. Based on the staining results, follicles were manually counted, and the area of ​​mouse ovarian fibrosis was calculated.

[0061] The results are shown in the table below. Figure 1-4 As shown.

[0062] Table 1. Estrogenic cycle of 8-month-old mice

[0063] In this context, 1 represents proestrus, 2 represents estrus, and 3 represents meestrus or interestrus.

[0064] Figure 1 The results show the follicle counts of eight-month-old mice one month after treatment with adenosine. It can be seen that after one month of daily injection of adenosine solution, the ovarian reserve of the mice was significantly restored. Compared with the control group, the number of primordial follicles (ovarian reserve) in the treatment group increased by 118.36%, without affecting the production of other grades of follicles.

[0065] Figure 2 The results of the mouse estrous cycle statistics are presented in Table 1. It can be seen that after one month of daily injection of adenosine solution, the number of aged mice with normal estrous cycles increased; compared to the control group, the number of aged mice with normal estrous cycles in the treatment group increased by 23.07%.

[0066] Figure 3The results show the ovarian area of ​​eight-month-old mice after one month of adenosine administration. It can be seen that daily injection of adenosine solution for one month increased the ovarian area of ​​the mice. Compared with the control group, the ovarian area of ​​the treated mice increased by 42.6%.

[0067] Figure 4 Figure A shows the statistical results of ovarian fibrosis area in eight-month-old mice after one month of adenosine administration. It can be seen that after one month of daily injection of adenosine solution, the ovarian fibrosis area in mice was significantly reduced. Compared with the control group, the ovarian fibrosis area in the treatment group was reduced by 45.35%.

[0068] Figure 4 Figure B shows the statistical results of ovarian fibrosis area in 12-month-old mice after two months of adenosine administration. It can be seen that after two months of daily injection of adenosine solution, the ovarian fibrosis area in mice was significantly reduced. Compared with the control group, the ovarian fibrosis area in the treatment group was reduced by 69.34%.

[0069] The results of experiments using adenosine analogues were similar to those of adenosine administration.

[0070] In summary, through statistical analysis of follicle count, ovarian area, ovarian fibrosis area, and estrous cycle, it can be determined that administration of adenosine or its analogues resulted in the recovery of primordial follicles, a reduction in fibrosis area, and an improvement in estrous cycle in mice. Therefore, it can be used to restore ovarian function and slow down ovarian fibrosis.

[0071] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. The use of compounds with the structure shown in Formula I in the preparation of medicaments for the prevention and / or treatment of ovarian fibrosis. in, R is selected from O or CH2.

2. Application of compounds with the structure shown in Formula I in the preparation of aphrodisiac drugs; in, R is selected from O or CH2.

3. The application of compounds with the structure shown in Formula I in the preparation of drugs that enhance or improve female reproductive capacity; in, R is selected from O or CH2.

4. The application according to claim 3, characterized in that, The improvement or enhancement of female fertility includes: Enhance ovarian reserve and increase ovarian area by at least one of the following:

5. The application according to any one of claims 1-3, characterized in that, The compound is adenosine or a pharmaceutically acceptable salt thereof, or mongolicin or a pharmaceutically acceptable salt thereof.

6. The application according to any one of claims 1-3, characterized in that, The drug also contains pharmaceutically acceptable excipients.

7. The application according to claim 6, characterized in that, The excipients include at least one of diluents, absorbents, binders, disintegrants, lubricants, colorants, coating materials, solvents, pH adjusters, antibacterial agents, isotonic adjusters, and chelating agents.

8. The application according to any one of claims 1-3, characterized in that, The drug also contains a second active substance; wherein the second active substance has at least one of the following functions (1)-(5): (1) Improve or restore ovarian function; (2) To treat or improve menstrual disorders; (3) Increase or improve estrogen (E2) levels; (4) Increase or improve anti-Müllerian hormone (AMH) levels; (5) Improve fertilization rate, high-quality embryo rate or egg retrieval rate.

9. A method for improving fertility, comprising: Administer an effective amount of the compound with the structural formula shown in Formula I to the subject; R is selected from O or CH2.

10. The method according to claim 9, characterized in that, The subjects were either normal individuals or patients with reproductive defects.

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