Anti-aging use of clomiphene and its derivatives

Abstract

The application provides anti-aging use of cromolyn and derivatives thereof. Specifically, the application provides use of cromolyn or a kind of cromolyn N-ethyl-N-phenyl acrylamide derivative in preparation of an anti-aging composition or preparation. The cromolyn and derivatives thereof of the application can prolong the lifespan of nematodes; the derivative JM03 can also significantly prolong the antioxidant stress resistance and osmotic stress resistance of nematodes. Therefore, the cromolyn and derivatives thereof of the application have application prospects in the aspect of anti-aging.

Description

Technical Field

[0001] This invention relates to the pharmaceutical field. Specifically, it relates to the anti-aging uses of clomiphene and its derivatives. Background Technology

[0002] Aging is a spontaneous and inevitable process that occurs in living organisms over time. It is primarily characterized by a decline in adaptability and resistance, impaired physiological functions, degenerative changes in tissues and organs, and ultimately, inevitable death. Accompanying aging, the incidence of many chronic diseases, such as heart disease, cancer, diabetes, and Alzheimer's disease, increases dramatically. In recent years, with the increase in average life expectancy and the decline in birth rates, the global population age structure has been constantly changing, primarily manifested in a continuous decrease in the number of people aged 0-15 years, while the number of people aged 65 and above has been continuously increasing. Currently, the number of people aged 65 and above is significantly higher than the number of people under 5 years old, and this trend will continue, meaning that the global aging population is intensifying. This demographic change will have a significant impact on the operation of national healthcare systems and place a heavy burden on economic development. Therefore, delaying aging and preventing, delaying, mitigating, or even reversing the progression of age-related diseases have become key to solving the aging problem for countries worldwide, and the search for anti-aging drugs is urgent.

[0003] However, there is currently a lack of satisfactory anti-aging drugs. Therefore, there is an urgent need in the field to develop new, safe and effective anti-aging methods and pharmaceutical compositions. Summary of the Invention

[0004] The purpose of this invention is to provide a safe and effective method and pharmaceutical composition for anti-aging. Specifically, this invention provides anti-aging uses for clomiphene and its derivatives JM01, JM02, JM03, JM04, JM05, JM06, JM07, JM08, JM09, JM10, JM11, JM12, JM13, and JM14.

[0005] In a first aspect of the invention, there is provided the use of an active ingredient or a composition containing said active ingredient in the preparation of an anti-aging pharmaceutical composition, characterized in that said active ingredient is cromatathan, or a derivative thereof, or a pharmaceutically acceptable salt thereof.

[0006] In another preferred embodiment, the clomiton derivative is of the type shown in formula (I). N -Ethyl- N -Phenylacetamide derivatives, or their optical isomers, or their racemates, or their pharmaceutically acceptable salts, or their solvates, or their hydrates, or their pharmaceutically acceptable prodrug esters:

[0007]

[0008] Formula (I)

[0009] In formula (I),

[0010] n is a positive integer selected from 0, 1, 2, 3, 4, 5, preferably n is 1, 2, or 3.

[0011] In the formula, R 1 R 1 and R 3 Each group is independently selected from: hydrogen, fluorine, chlorine, bromine, carboxyl, aryl, substituted aryl, C 1-6 Alkyl, substituted C 1-6 Alkyl, C3-C6 cycloalkyl, substituted C3-C6 cycloalkyl, C 1-6 Acyl group, substituted acyl group, C 2-7 Ester group, substituted C 2-7 Ester group.

[0012] In another preferred embodiment, the carboxyl group is COOH.

[0013] In another preferred embodiment, the substitution refers to having one or more substituents selected from the group consisting of: C1-C4 alkyl, C3-C6 cycloalkyl, halogen, benzene ring, heterocycle, five-membered ring, and six-membered ring.

[0014] In another preferred embodiment, the aryl group is a C6-C10 aryl group.

[0015] In another preferred embodiment, the ester group is -COOR. 4 , where R 4 It is a C1-C4 alkyl group.

[0016] In another preferred embodiment, R1 is selected from the group consisting of H, C1-C4 alkyl, halogen, carboxyl, or combinations thereof.

[0017] In another preferred embodiment, R2 is selected from the group consisting of H, C1-C4 alkyl, halogen, carboxyl, ester, or combinations thereof.

[0018] In another preferred embodiment, R3 is selected from the group consisting of: H, C1-C4 alkyl, and C3-C6 cycloalkyl.

[0019] In another preferred embodiment, the crometton and its N -Ethyl- N -Phenylacetamide derivatives are selected from the following group:

[0020]

[0021] In another preferred embodiment, the pharmaceutical composition is used for one or more applications selected from the group consisting of:

[0022] i) Extend lifespan and delay aging;

[0023] ii) Increase antioxidant capacity;

[0024] iii) Increase resistance to osmotic stress;

[0025] iv) Improve health physiological parameters and reduce the incidence of age-related diseases.

[0026] In another preferred embodiment, the composition is a pharmaceutical composition or a cosmetic composition.

[0027] In another preferred embodiment, the pharmaceutical composition contains (a) clomiphene, or thereof N -Ethyl- N (a) a phenylacrylamide derivative or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or excipient.

[0028] In another preferred embodiment, the pharmaceutical composition further includes additional anti-aging active ingredients.

[0029] In another preferred embodiment, the active ingredient in the pharmaceutical composition is present in an amount of 0.001-99 wt%, more preferably 0.01-90 wt%, based on the total weight of the composition.

[0030] In another preferred embodiment, the pharmaceutical composition is an injectable or oral medication.

[0031] In another preferred embodiment, the pharmaceutical composition is selected from the group consisting of tablets, capsules, granules, suspensions, pills, solutions, syrups, or injections.

[0032] In another preferred embodiment, the pharmaceutical composition is applied to mammals or nematodes.

[0033] In another preferred embodiment, the mammal includes primates; more preferably, the mammal is a human.

[0034] In another preferred embodiment, the anti-aging includes skin anti-aging.

[0035] In a second aspect of the invention, a pharmaceutical composition for anti-aging is provided, characterized in that the pharmaceutical composition comprises:

[0036] (a1) Cromiton, or its derivatives N -Ethyl- N 1-Phenylacetamide derivatives or their pharmaceutically acceptable salts may be used as active ingredients;

[0037] (a2) Additional anti-aging active ingredients, wherein the additional anti-aging active ingredients are selected from the group consisting of: metformin, resveratrol, rapamycin, acarbose, dasatinib, quercetin, curcumin, astaxanthin, and arbutin; and

[0038] (b) Pharmaceutically acceptable carriers or excipients.

[0039] In another preferred embodiment, the content of the active ingredient in the pharmaceutical composition (a1) is 0.001-99 wt%, preferably 0.01-90 wt%, based on the total weight of the composition.

[0040] In another preferred embodiment, the pharmaceutical composition is an injectable or oral medication.

[0041] In another preferred embodiment, the pharmaceutical composition is selected from the group consisting of tablets, capsules, granules, suspensions, pills, solutions, syrups, or injections.

[0042] In a third aspect of the invention, a cosmetic composition or product is provided, characterized in that the cosmetic composition or product contains cromite, or thereof. N -Ethyl- N -Phenylacetamide derivatives, or pharmaceutically acceptable salts thereof, are used as effective anti-aging ingredients.

[0043] In another preferred embodiment, the content of the active ingredient in the cosmetic composition or product is 0.001-99 wt%, more preferably 0.01-90 wt%, based on the total weight of the composition.

[0044] In another preferred embodiment, the cosmetic composition or product further includes additional components selected from the group consisting of whitening or spot-reducing ingredients, anti-inflammatory ingredients, antioxidant ingredients, UV-protective ingredients, or combinations thereof.

[0045] In another preferred embodiment, the dosage form of the cosmetic composition or product is selected from the group consisting of solid dosage forms, semi-solid dosage forms, liquid dosage forms, or combinations thereof.

[0046] In another preferred embodiment, the cosmetic product is prepared in the form selected from the group consisting of: solution, gel, cream, lotion, serum, transdermal patch, mask, or a combination thereof.

[0047] In a fourth aspect of the invention, an anti-aging method is provided, characterized by comprising the steps of: administering to a desired subject a) a pharmaceutical composition containing clomiphene or a clomiphene derivative, or a pharmaceutically acceptable salt thereof; b) the pharmaceutical composition of the second aspect of the invention; or c) the cosmetic composition or product of the third aspect of the invention.

[0048] In another preferred embodiment, the object includes mammals or nematodes.

[0049] In another preferred embodiment, the object includes primates; more preferably, the object is a human.

[0050] In another preferred embodiment, the method of administration is selected from the group consisting of: oral administration, injection administration, topical administration, application of cosmetics to the skin, or a combination thereof.

[0051] It should be understood that, within the scope of this invention, the above-described technical features of this invention and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here. Attached Figure Description

[0052] The following figures are used to illustrate specific embodiments of the present invention and are not intended to limit the scope of the invention as defined by the claims.

[0053] Figure 1 The results showed that clomiphene and its derivatives JM01, JM02, JM03, JM04, JM05, JM06, JM07, and JM08 could prolong the lifespan of nematodes at concentrations of 100 μM and 400 μM.

[0054] Figure 2 The results showed that clomiton derivatives JM09, JM10, JM11, JM12, JM13, and JM14 could prolong the lifespan of nematodes at concentrations of 100 μM and 400 μM.

[0055] Figure 3 The results showed that the clomiphene derivative JM03 at a concentration of 400 μM could significantly increase the antioxidant stress resistance of nematodes.

[0056] Figure 4 The results showed that the clomiton derivative JM03 at a concentration of 400 μM could significantly increase the osmotic stress resistance of nematodes. Detailed Implementation

[0057] Through extensive and in-depth research, the inventors unexpectedly discovered for the first time that clomiphene and its derivatives possess the ability to prolong lifespan, delay aging, and increase resistance to oxidative stress and osmotic stress. This led to the discovery that clomiphene can be applied to anti-aging, improving healthy physiological parameters, and reducing the incidence of age-related diseases. Based on this, the present invention was completed.

[0058] Specifically, in this invention, nematodes were used as model animals, and experiments on nematode lifespan confirmed that the clomiphene and its derivatives described in this invention can prolong the lifespan of nematodes; further experiments also confirmed that its derivative JM03 can significantly prolong the nematode's ability to resist oxidative stress and osmotic stress.

[0059] Aging and Anti-aging

[0060] As used herein, "aging" refers to the loss and degeneration of an organism's constituent substances, tissue structure, and physiological functions over time. In this invention, aging refers to biological aging.

[0061] As used in this article, "anti-aging" refers to delaying, blocking, reducing, stopping, and / or reversing the effects or process of aging.

[0062] Cellular senescence causes individual cells to cease dividing, eventually stopping after a few divisions. Senescent cells can interfere with vital functions throughout the organism and lead to various disorders. The aging of the entire organism is accompanied by an increased risk of certain disorders, such as diseases, complications, and symptoms.

[0063] Caenorhabditis elegans

[0064] Caenorhabditis elegans is a classic model organism, characterized by its short lifespan, ease of observation, and clear genetic background. Furthermore, its genes are highly homologous to many important human functional genes and share similar signal transduction pathways, making it increasingly important in pathological research, novel target discovery, and drug screening.

[0065] Caenorhabditis elegans is an important model organism for anti-aging research and has been used for screening anti-aging substances and studying the mechanisms of anti-aging.

[0066] Active ingredients

[0067] As used herein, the terms “active ingredient of the present invention,” “anti-aging active ingredient of the present invention,” “compound of the present invention,” and “anti-aging compound of the present invention” refer to clomiphene or clomiphene derivatives, or pharmaceutically acceptable salts thereof, or solvates thereof, or optical isomers thereof, or racemates thereof, or crystals thereof, or hydrates thereof, or deuterated compounds thereof, or combinations thereof.

[0068] Cromitrin is an old anti-scabies drug that can also be used to treat pubic lice and other skin conditions such as pruritus. Its chemical formula is:

[0069]

[0070] In another preferred embodiment, the clomiton derivative is of the type shown in formula (I). N -Ethyl-N -Phenylacetamide derivatives, or their optical isomers, or their pharmaceutically acceptable salts, hydrates, or their pharmaceutically acceptable prodrug esters:

[0071]

[0072] Formula (I)

[0073] In the formula, R 1 ~R 3 They are: hydrogen, fluorine, chlorine, bromine, carboxyl, aryl, C 1-6 Alkyl, C 1-6 Acyl group.

[0074] In another preferred embodiment, the crometton N -Ethyl- N -Phenylacetamide derivatives are selected from the following group:

[0075]

[0076] Through research, the inventors have discovered that the active ingredients of this invention have the ability to significantly prolong lifespan, delay aging, and increase resistance to oxidative stress and osmotic stress. They are expected to be developed into anti-aging drugs and can be used to prolong lifespan, delay aging, and prepare anti-aging drugs, cosmetics, etc.

[0077] Pharmaceutical Compositions and Administration

[0078] The present invention also provides pharmaceutical compositions containing the active ingredients of the present invention, said pharmaceutical compositions being used for anti-aging.

[0079] A preferred composition is a pharmaceutical composition containing an effective amount of clomiphene, a clomiphene derivative thereof, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

[0080] As used herein, the term “effective amount” or “effective dose” refers to an amount that is functional or active (i.e., anti-aging) in humans and / or animals and is acceptable to humans and / or animals.

[0081] As used herein, the term "pharmaceuticalally acceptable" refers to a substance suitable for human and / or mammalian use without excessive adverse side effects (such as toxicity, irritation, and allergic reactions), i.e., a substance with a reasonable benefit / risk ratio. The term "pharmaceuticalally acceptable carrier" refers to a carrier used for the administration of a therapeutic agent, including various excipients and diluents.

[0082] The pharmaceutical compositions of the present invention contain a safe and effective amount of the active ingredient of the present invention. The anti-aging active ingredient in the pharmaceutical composition includes clomiphene and its derivatives JM01, JM02, JM03, JM04, JM05, JM06, JM07, JM08, JM09, JM10, JM11, JM12, JM13, and JM14. Further, the pharmaceutical composition also contains a pharmaceutically acceptable carrier or excipient. Carriers include (but are not limited to): saline, buffer solution, glucose, water, glycerol, ethanol, and combinations thereof. The pharmaceutical composition is for injection or oral administration. Generally, the pharmaceutical composition should be matched to the route of administration. The dosage form of the pharmaceutical compositions of the present invention is selected from tablets, capsules, oral liquids, granules, suspensions, pills, solutions, syrups, or injections. For example, it is prepared using physiological saline or an aqueous solution containing glucose and other excipients by conventional methods. The pharmaceutical composition is preferably manufactured under aseptic conditions.

[0083] The effective amount of the active ingredient described in this invention can vary depending on the administration method and the severity of the disease to be treated. A preferred effective amount can be determined by those skilled in the art based on various factors (e.g., through clinical trials). These factors include, but are not limited to: pharmacokinetic parameters of the active ingredient, such as bioavailability, metabolism, and half-life; the severity of the disease to be treated, the patient's weight, the patient's immune status, and the route of administration. Generally, satisfactory results are obtained when the active ingredient of this invention is administered daily at a dose of approximately 0.00001 mg to 50 mg / kg animal body weight (preferably 0.0001 mg to 10 mg / kg animal body weight). For example, due to the urgency of the treatment condition, several separate doses may be administered daily, or the dose may be reduced proportionally.

[0084] Typically, when clomiphene is administered orally, the average daily dose for a 60 kg subject is usually 10-500 mg, preferably 20-300 mg, and more preferably 50-250 mg. The daily dose may be divided into one, two, or multiple administrations.

[0085] The pharmaceutically acceptable carriers described in this invention include (but are not limited to): water, saline, liposomes, lipids, peptides, cellulose, nanogels, or combinations thereof. The choice of carrier should be matched to the route of administration, as is well known to those skilled in the art.

[0086] Anti-aging methods

[0087] This invention provides an anti-aging method (including therapeutic methods or in vitro non-therapeutic methods).

[0088] Typically, the method includes the step of administering clomiphene, or its N-ethyl-N-phenylacrylamide derivative, or a pharmaceutically acceptable salt thereof, to the desired subject as an anti-aging active ingredient.

[0089] Preferably, the anti-aging method involves applying (or coating) a cosmetic composition or product to the skin of the subject, wherein the cosmetic composition or product contains cromite or its derivatives as an effective anti-aging ingredient.

[0090] The effective anti-aging ingredients in the cosmetics include clomiphene and its derivatives JM01, JM02, JM03, JM04, JM05, JM06, JM07, JM08, JM09, JM10, JM11, JM12, JM13, and JM14.

[0091] In this invention, the dosage form of cosmetics may be selected from the following group (but is not limited to): solid dosage form, semi-solid dosage form, liquid dosage form, or a combination thereof.

[0092] Typically, the cosmetic products of the present invention are prepared in the form of a solution, gel, cream, lotion, serum, transdermal patch, mask, or a combination thereof.

[0093] The main advantages of this invention include:

[0094] 1) It was the first time that clomiphene was confirmed to be effective in anti-aging and to significantly extend the lifespan of organisms or cells.

[0095] 2) Clomidon can also improve the body's health, such as enhancing its ability to resist oxidative stress and osmotic stress.

[0096] 3) Clomidon, as a marketed drug, has a good safety profile.

[0097] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed under conventional conditions, such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or as recommended by the manufacturer. Unless otherwise stated, percentages and parts are weight percentages and parts by weight.

[0098] Example 1: Preparation of Clomidon Derivatives

[0099] The synthetic route for clomiton derivatives JM01-JM14 is shown in Flowchart 1, where the reagents and reaction conditions are: (a) Acryloyl chloride derivatives, K2CO3, CH2Cl2, 0 °C to rt; (b) 1M NaOH (aq.), CH3OH, rt. Using substituted N-ethylaniline derivatives as starting materials, a condensation reaction was carried out with acryloyl chloride derivatives under alkaline conditions of potassium carbonate, yielding JM01-JM05, JM10, JM12, JM13 and intermediates 9-11 in 90-98% yield. Subsequently, the ester groups of compounds JM10, JM12, and intermediates 9-11 were hydrolyzed with 1M sodium hydroxide aqueous solution to give the final products JM06-JM09, JM11, and JM14.

[0100]

[0101] All reagents used in the chemical synthesis were purchased from commercial companies and used directly. Proton and carbon NMR spectra were performed using Bruker 400 MHz and 600 MHz NMR spectra at 303 K, with tetramethylsilane (TMS) as the internal standard. Chemical shifts (δ) are expressed in ppm. High-resolution mass spectrometry was performed on a Waters LCT or Agilent 6545 Q-TOF. Column chromatography was performed using 200-300 mesh silica gel, monitored at wavelengths of 254 nm and 365 nm.

[0102] General synthetic routes for compounds JM01-JM05, JM10, JM12, JM13, and intermediates 9-11

[0103] Compounds 1–8 (4.0 mmol) were dissolved in dichloromethane, and then an acryloyl chloride derivative (4.0 mmol) and potassium carbonate (1.66 g, 12.0 mmol) were added at 0 °C. After stirring at room temperature for 1 h, the solvent was evaporated to dryness under reduced pressure. The residue was dissolved in ethyl acetate and extracted sequentially with water and saturated brine. The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness, and purified by column chromatography to obtain the target product.

[0104] ( E )- N -Ethyl- N -(3-Methylphenyl)-2-butenamide (JM01)

[0105]

[0106] 780 mg, 96.0% yield. 1 H NMR (600 MHz, CDCl3) d7.33–7.27 (m, 1H), 7.16 (d, J = 7.5 Hz, 1H), 6.97 (s, 1H), 6.96 – 6.87 (m, 2H), 5.68 (d, J =14.8 Hz, 1H), 3.80 (q, J = 6.9 Hz, 2H), 2.39 (s, 3H), 1.72 (d, J = 6.7 Hz, 3H), 1.13 (t, J = 7.0 Hz, 3H). ESI-HRMS [M+H] + calcd for C 13 H 18 NO: 204.1383, found: 204.1389.

[0107] ( E )- N -Ethyl- N -(4-Methylphenyl)-2-butenamide (JM02)

[0108]

[0109] 789 mg, 97.0% yield. 1 H NMR (600 MHz, CDCl3) d 7.21 (d, J = 7.5 Hz, 2H), 7.03 (d, J = 7.4 Hz, 2H), 6.94 – 6.86 (m, 1H), 5.68 (d, J = 15.0 Hz, 1H), 3.79 (q, J = 7.0 Hz, 2H), 2.39 (s, 3H), 1.71 (d, J = 6.7 Hz, 3H), 1.12(t, J = 7.0 Hz, 3H). ESI-HRMS [M+H] + calcd for C 13 H 18 NO: 204.1383, found: 204.1386.

[0110] ( E )- N -Ethyl- N-(2-Fluorophenyl)-2-butenamide (JM03)

[0111]

[0112] 791 mg, 95.4% yield. 1 H NMR (600 MHz, CDCl3) d 7.39 – 7.34 (m, 1H),7.23 – 7.16 (m, 3H), 6.99-6.91 (m, 1H), 5.63 (d, J = 15.0 Hz, 1H), 3.87-3.81(m, 1H), 3.78 – 3.69 (m, 1H), 1.73 (dd, J = 6.9, 1.5 Hz, 3H), 1.12 (t, J =7.2 Hz, 3H). ESI-HRMS [M+H] + calcd for C 12 H 15 FNO: 208.1132, found: 208.1135.

[0113] ( E )- N -Ethyl- N -(2-Chlorophenyl)-2-butenamide (JM04)

[0114]

[0115] 828 mg, 92.5% yield. 1 H NMR (600 MHz, CDCl3) d 7.54 – 7.50 (m, 1H),7.37 – 7.32 (m, 2H), 7.24 – 7.20 (m, 1H), 6.97-6.91 (dq, J = 13.9, 6.9 Hz,1H), 5.53 – 5.46 (m, 1H), 4.07 (dq, J = 14.3, 7.2 Hz, 1H), 3.48 (dq, J =14.3, 7.2 Hz, 1H), 1.72 (dd, J = 6.9, 1.6 Hz, 3H), 1.14 (t, J= 7.2 Hz, 3H). ESI-HRMS [M+H] + calcd for C 12 H 15 ClNO: 224.0837, found: 224.0804.

[0116] ( E )- N -Ethyl- N -(2-Bromophenyl)-2-butenamide (JM05)

[0117]

[0118] 1009 mg, 94.1% yield. 1 H NMR (600 MHz, CDCl3) d 7.70 (dd, J = 8.0, 1.3 Hz, 1H), 7.39 (td, J = 7.6, 1.3 Hz, 1H), 7.28 – 7.25 (m, 1H), 7.22 (dd, J = 7.8, 1.6 Hz, 1H), 6.95 (dq, J = 13.9, 6.9 Hz, 1H), 5.48 (dd, J = 15.0, 1.6Hz, 1H), 4.14 (dq, J = 14.3, 7.2 Hz, 1H), 3.39 (dq, J = 14.2, 7.2 Hz, 1H), 1.72 (dd, J = 6.9, 1.6 Hz, 3H), 1.15 (t, J = 7.2 Hz, 3H). ESI-HRMS [M+H] + calcd for C 12 H 15 BrNO: 268.0332, found: 268.0308.

[0119] ( E )- N -Ethyl- N -(4-Methylphenyl)fumaric acid monoethyl ester amide (JM10)

[0120]

[0121] 951 mg, 91.0% yield. 1 H NMR (600 MHz, CDCl3) d 7.22 (d, J = 8.0 Hz, 2H), 7.02 (d, J = 8.1 Hz, 2H), 6.82 (q, J = 15.3 Hz, 2H), 4.16 (q, J = 7.1Hz, 2H), 3.83 (q, J = 7.1 Hz, 2H), 2.39 (s, 3H), 1.24 (t, J = 7.1 Hz, 3H), 1.15 (t, J = 7.1 Hz, 3H). 13 C NMR (150 MHz, CDCl3) d 165.81, 163.59, 138.29,138.26, 134.83, 130.71, 130.42, 127.84, 60.92, 44.66, 21.10, 14.10, 12.81.ESI-HRMS [M+H] + calcd for C 15 H 20 NO3: 262.1438, found: 262.1435.

[0122] ( E )- N -Ethyl- N -(2-Fluorophenyl)fumaric acid monoethyl ester amide (JM12)

[0123]

[0124] 960 mg, 90.5% yield. 1 H NMR (600 MHz, CDCl3) d 7.43-7.37 (m, 1H),7.25 – 7.17 (m, 3H), 6.87 (d, J = 15.2 Hz, 1H), 6.74 (d, J = 15.2 Hz, 1H), 4.16 (q, J = 7.1 Hz, 2H), 3.90 – 3.77 (m, 2H), 1.25 (t,J = 7.1 Hz, 3H), 1.15(t, J = 7.1 Hz, 3H). ESI-HRMS [M+H] + calcd for C 14 H 17 FNO3: 266.1187, found:266.1185.

[0125] ( E )- N -Ethyl- N -(4-Fluorophenyl)-2-butenamide (JM13)

[0126]

[0127] 792 mg, 95.5% yield. 1 H NMR (600 MHz, CDCl3) d 7.15 – 7.09 (m, 4H), 6.92 (dq, J = 14.0, 6.8 Hz, 1H), 5.62 (d, J = 14.9 Hz, 1H), 3.78 (q, J = 7.1Hz, 2H), 1.73 (d, J = 6.8 Hz, 3H), 1.13 (t, J = 7.1 Hz, 3H). ESI-HRMS [M+H] + calcd for C 12 H 15 FNO: 208.1132, found: 208.1132.

[0128] General synthetic routes for compounds JM06-JM09, JM11, and JM14

[0129] Compounds JM10, JM12, and intermediate 9-11 (2.0 mmol) were dissolved in methanol, and 5 mL of 1 M sodium hydroxide was added. The mixture was stirred at room temperature for 2 h. The methanol solvent was evaporated to dryness under reduced pressure, and the pH was adjusted to less than 2 with 1 M dilute hydrochloric acid aqueous solution. The mixture was extracted with ethyl acetate, and the combined organic phases were dried over anhydrous sodium sulfate. After concentration under reduced pressure, the product was purified by column chromatography to obtain the target product.

[0130] ( E )- N -Ethyl- N -(2-Methyl-5-carboxyphenyl)-2-butenamide (JM06)

[0131]

[0132] 485 mg, 98.0% yield. 1 H NMR (600 MHz, CDCl3) d 10.91 (s, 1H), 8.04 (dd, J = 8.0, 1.3 Hz, 1H), 7.86 (d, J = 1.3 Hz, 1H), 7.43 (d, J = 8.0 Hz, 1H), 7.01 (dq, J = 13.9, 6.9 Hz, 1H), 5.50 (dd, J = 15.0, 1.4 Hz, 1H), 4.11(dq, J = 14.2, 7.2 Hz, 1H), 3.45 (dq, J = 14.2, 7.2 Hz, 1H), 2.29 (s, 3H), 1.72 (dd, J = 6.9, 1.2 Hz, 3H), 1.18 (t, J = 7.2 Hz, 3H). ESI-HRMS [M+H] + calcd for C 14 H 18 NO3: 248.1281, found: 248.1281.

[0133] ( E )- N -Ethyl- N -(2-Methyl-4-carboxyphenyl)-2-butenamide (JM07)

[0134]

[0135] 478 mg, 97.0% yield. 1 H NMR (400 MHz, CDCl3) d 8.09 (d, J = 1.3 Hz, 1H), 8.02 (d, J = 8.0 Hz, 1H), 7.22 (d, J = 8.1 Hz, 1H), 7.00 (dq, J = 13.8, 6.8 Hz, 1H), 5.49 (d, J= 14.9 Hz, 1H), 4.09 (dq, J = 14.0, 7.1 Hz, 1H), 3.46(dq, J = 13.9, 7.0 Hz, 1H), 2.28 (s, 3H), 1.72 (d, J = 6.8 Hz, 3H), 1.17 (t, J = 7.1 Hz, 3H). ESI-HRMS [M+H] + calcd for C 13 H 18 NO: 248.1281, found: 248.1281.

[0136] ( E )- N -Ethyl- N -(2-Methylphenyl)fumaramide (JM08)

[0137]

[0138] 460 mg, 98.5% yield. 1 H NMR (600 MHz, CDCl3) d 7.33 – 7.29 (m, 2H),7.27 – 7.23 (m, 1H), 7.07 (d, J = 7.6 Hz, 1H), 6.83 (d, J = 15.3 Hz, 1H), 6.65 (d, J = 15.3 Hz, 1H), 4.14 – 4.06 (m, 1H), 3.47 – 3.40 (m, 1H), 2.18 (s,3H), 1.16 (t, J = 7.2 Hz, 3H). ESI-HRMS [M+H] + calcd for C 13 H 16 NO3: 234.1125, found: 234.1122.

[0139] ( E )- N -Ethyl- N -(4-Methylphenyl)fumaramide (JM09)

[0140]

[0141] 464 mg, 99.5% yield. 1 H NMR (600 MHz, CDCl3) d 7.22 (d, J = 7.7 Hz, 2H), 7.00 (d, J = 7.6 Hz, 2H), 6.80 (s, 2H), 3.83 (q, J = 7.1 Hz, 2H), 2.39(s, 3H), 1.14 (t, J = 7.1 Hz, 3H). 13 C NMR (150 MHz, CDCl3) d 169.96, 163.42,138.49, 138.02, 136.52, 130.48, 129.85, 127.75, 44.79, 21.12, 12.75. ESI-HRMS[M+H] + calcd for C 13 H 16 NO3 + : 234.1125, found: 234.1129.

[0142] ( E )- N -Ethyl- N -(2-Fluorophenyl)fumaramide (JM11)

[0143]

[0144] 464 mg, 97.8% yield. 1 H NMR (600 MHz, CDCl3) d 7.43 – 7.38 (m, 1H),7.24-7.16 (m, 3H), 6.83 (d, J = 15.2 Hz, 1H), 6.77 (d, J = 15.2 Hz, 1H), 3.89– 3.76 (m, 2H), 1.15 (t, J = 7.2 Hz, 3H). ESI-HRMS [M+H] + calcd for C 12 H 13 FNO3:238.0874, found: 238.0874.

[0145] ( E )- N -Ethyl- N -(4-Fluorophenyl)fumaramide (JM14)

[0146]

[0147] 471 mg, 99.2% yield. 1 H NMR (600 MHz, CDCl3) d 7.13 (d, J = 6.2 Hz, 4H), 6.79 (q, J = 15.3 Hz, 2H), 3.83 (q, J = 6.9 Hz, 2H), 1.15 (t, J = 7.0Hz, 3H). ESI-HRMS [MH] - calcd for C 12 H 11 FNO3 - : 236.0728, found: 236.0724.

[0148] Example 2: Nematode Lifespan Experiment

[0149] Clomid was purchased from Shanghai Maclean Biotechnology Co., Ltd.; wild-type Caenorhabditis elegans (N2) was purchased from Caenorhabditis Genetics Center (CGC). Synchronized nematodes were transferred at stage L4 to nematode growth medium (NGM) containing a blank control group and clomid and its derivatives (100 / 400 μM). The medium was changed every other day, and the nematodes were cultured at 20°C. The number of surviving nematodes was recorded daily, and dead nematodes were removed from the culture dish until all nematodes died. Data were summarized, survival curves were plotted using Graphpad Prism 8.3.0, and statistical significance (Log-rank test) was calculated.

[0150] Results: Lifetime experiment data are shown in Table 1. Figure 1 and Figure 2 As shown,

[0151] Table 1. Lifetime experimental data of crometton and its derivatives

[0152]

[0153] Figure 1As shown in this embodiment, clomiphene and its derivatives JM01, JM02, JM03, JM04, JM05, JM06, JM07, and JM08 can prolong the lifespan of nematodes at concentrations of 100 μM and 400 μM. Among them, clomiphene citrate extended the lifespan of nematodes by 6.43% at a concentration of 400 μM; JM01 extended the lifespan by 23.20% at a concentration of 400 μM; JM02 extended the lifespan by 31.42% and 16.51% at concentrations of 100 μM and 400 μM, respectively; JM03 extended the lifespan by 25.66% and 19.39% at concentrations of 100 μM and 400 μM, respectively; JM04 extended the lifespan by 25.25% and 20.64% at concentrations of 100 μM and 400 μM, respectively; JM05 extended the lifespan by 16.13% and 25.55% at concentrations of 100 μM and 400 μM, respectively; JM06 extended the lifespan by 6.86% and 9.49% at concentrations of 100 μM and 400 μM, respectively; and JM07 extended the lifespan by 6.43% at a concentration of 400 μM and 400 μM, respectively. At a concentration of μM, JM08 extended the lifespan of nematodes by 7.74%; at concentrations of 100 μM and 400 μM, JM08 extended the lifespan of nematodes by 12.75% and 16.51%, respectively.

[0154] Figure 2 As shown in this embodiment, clomiphene and its derivatives JM09, JM10, JM11, JM12, JM13, and JM14 can prolong the lifespan of nematodes at concentrations of 100 μM and 400 μM. Among them, JM09 extended the lifespan of nematodes by 10.84% ​​and 14.06% at concentrations of 100 μM and 400 μM, respectively; JM10 extended the lifespan of nematodes by 5.95% and 4.93% at concentrations of 100 μM and 400 μM, respectively; JM11 extended the lifespan of nematodes by 7.88% and 14.89% at concentrations of 100 μM and 400 μM, respectively; JM12 extended the lifespan of nematodes by 8.99% at a concentration of 400 μM; JM03 extended the lifespan of nematodes by 6.13% and 11.49% at concentrations of 100 μM and 400 μM, respectively; and JM14 extended the lifespan of nematodes by 3.27% and 8.71% at concentrations of 100 μM and 400 μM, respectively.

[0155] This example illustrates that clomiphene and its derivatives can significantly extend the lifespan of nematodes.

[0156] Example 3: Nematode Antioxidant Stress Experiment

[0157] Synchronized nematodes were transferred at stage L4 to the blank control group and the JM03 group (400 μM) nematode growth medium (NGM) containing FUDR, and cultured at 20°C until day 4 of adulthood. The nematodes were then transferred to 800 μL of M9 solution containing 10 mM paraquat. The experiment was conducted in 24-well plates, with 6-8 nematodes per well. The number of surviving nematodes was recorded every 2-3 h; nematodes that did not move upon contact with the platinum thread were considered dead. Data were summarized, survival curves were plotted using Graphpad Prism 8.3.0, and statistical significance was calculated using the Log-rank test.

[0158] Results: Experimental data on nematode antioxidant stress are as follows: Figure 3 As shown, JM03 at a concentration of 400 μM can significantly prolong the antioxidant stress resistance of nematodes.

[0159] Example 4: Nematode Osmotic Stress Experiment

[0160] Synchronized nematodes were transferred at stage L4 to nematode growth medium (NGM) containing FUDR in the blank control group and the JM03 group (400 μM). They were cultured at 20°C until day 4 of adulthood. The nematodes were washed with M9 solution and transferred to high-salt NGM dishes containing 500 mM NaCl. Excess M9 solution was blotted dry with filter paper. Timing was recorded from the point when the M9 solution was completely absorbed as 0 min. The number of crawling nematodes on the high-salt plate was then continuously recorded at 3 min, 5 min, 7 min, 9 min, 11 min, and 13 min until all nematodes were paralyzed. Survival curves were plotted after the experiment, and the statistical method used was Log-rank (Mantel Cox).

[0161] Results: Data from the nematode osmotic stress experiment are as follows: Figure 4 As shown, JM03 at a concentration of 400 μM can significantly prolong the osmotic stress resistance of nematodes.

[0162] All documents mentioned in this invention are incorporated herein by reference as if each document were individually incorporated by reference. Furthermore, it should be understood that after reading the foregoing teachings of this invention, those skilled in the art can make various alterations or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims.

Claims

1. Use of an active ingredient or a composition containing said active ingredient in the preparation of an anti-aging pharmaceutical composition, characterized in that, The active ingredients are selected from the following group:

2. The use as described in claim 1, characterized in that, The pharmaceutical composition is used to prolong life and delay aging.

3. The use as described in claim 1, characterized in that, The pharmaceutical composition is administered to mammals or nematodes.

4. The use as described in claim 3, characterized in that, The mammals mentioned include primates.

5. A pharmaceutical composition for anti-aging, characterized in that, The pharmaceutical composition contains: (a1) The following compounds are used as active ingredients; (a2) Additional anti-aging active ingredients, wherein the additional anti-aging active ingredients are selected from the group consisting of: metformin, resveratrol, rapamycin, acarbose, dasatinib, quercetin, curcumin, astaxanthin, and arbutin; and (b) Pharmaceutically acceptable carrier.

6. The pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition is an injectable or oral medication.

7. A cosmetic composition, characterized in that, The cosmetic composition contains compounds selected from the group consisting of or pharmaceutically acceptable salts thereof as active anti-aging ingredients:

8. The cosmetic composition according to claim 7, characterized in that, The cosmetic composition further includes additional components selected from the group consisting of: whitening or spot-reducing ingredients, anti-inflammatory ingredients, antioxidant ingredients, anti-UV ingredients, or combinations thereof.

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