Compound for improving mitochondrial function and preventing and treating aging and mitochondrial-related diseases, and preparation method therefor and use thereof

Abstract

The present application relates to the technical field of pharmaceutical chemistry, and provides a compound or a pharmaceutically acceptable salt thereof for improving mitochondrial function and preventing and treating aging and mitochondrial-related diseases, and a preparation method therefor and a use thereof. The compound has a structure as shown in formula (1). The compound provided in the present application has good solubility and high bioavailability, and can be used for preventing and / or treating diseases related to mitochondrial function.

Description

Compounds for improving mitochondrial function, preventing aging and mitochondrial-related diseases, their preparation methods and applications

[0001] This application claims priority to Chinese Patent Application No. CN202410936282.X, filed on July 12, 2024, entitled "Compounds for Improving Mitochondrial Function, Preventing Aging and Mitochondrial-Related Diseases, Preparation Methods and Applications thereof", the entire contents of which are incorporated herein by reference. Technical Field

[0002] This application relates to the field of medicinal chemistry, specifically to compounds for improving mitochondrial function, preventing and treating aging and mitochondrial-related diseases, as well as their preparation methods and applications. Background Technology

[0003] Mitochondria are energy-producing organelles within cells, primarily providing the energy cells need, namely nucleotide triphosphates (ATP). Mitochondrial diseases are a range of illnesses caused by abnormal mitochondrial function, involving neurodegenerative diseases, mental disorders, muscle diseases, and metabolic disorders. Some typical examples include Friedreich's ataxia, amyotrophic lateral sclerosis (ALS), spinocerebellar ataxia type 3, Duchenne muscular dystrophy, sarcopenia, and Pompe disease.

[0004] Friedreich ataxia (FA) is a progressive autosomal recessive neurodegenerative disorder. In the vast majority of patients, a biallelic trinucleotide (GAA) duplication is amplified in the first intron of the FXN gene, which impairs transcription and significantly reduces the amount of functional frataxin protein. Frataxin deficiency leads to a variety of pathological consequences, including interruption of iron-sulfur cluster biosynthesis, dysregulation of cellular iron, mitochondrial dysfunction, and increased sensitivity to in vitro oxidative stress, all of which ultimately contribute to the clinical symptoms of FA.

[0005] Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease of unknown etiology that primarily affects motor neurons in the cerebral cortex, brainstem, and spinal cord. Its main clinical manifestations include progressive skeletal muscle atrophy, weakness, fasciculations, bulbar palsy, and pyramidal tract signs. Studies show that the pathogenesis of ALS involves multiple pathological phenomena, including abnormal RNA processing, glutamate excitotoxicity, cytoskeleton disorganization, mitochondrial dysfunction, viral infection, apoptosis, abnormal growth factors, and inflammatory responses.

[0006] Spinocerebellar ataxia type 3 (SCA3), also known as Machado-Joseph disease (MJD), is a rare, inherited neurodegenerative disorder caused by an abnormal amplification of the CAG trinucleotide repeat in the AXTN3 gene coding sequence. Its main clinical symptoms include progressive cerebellar ataxia, pyramidal tract signs, rigid extrapyramidal syndrome, peripheral muscle atrophy and generalized reflex muscle dysfunction, and progressive extraocular muscle palsy. Currently, there is no cure for SCA3; existing treatments can only alleviate symptoms and slow disease progression.

[0007] Duchenne muscular dystrophy (DMD) is a serious, progressive, inherited muscle atrophy disorder that commonly affects male children. It is caused by a mutation in the gene encoding dystrophin. The mutation leads to a deficiency of myotrophin, which in turn promotes a pathological cascade involving structural defects in the muscle membrane, oxidative stress, mitochondrial dysfunction, chronic inflammation, muscle degeneration, and impaired regeneration, ultimately affecting the respiratory and cardiac muscles and leading to death.

[0008] Human aging involves a variety of changes, including gradual functional loss, decreased fertility, and increased mortality. Among all observations of humans (including health, nutrition, and physical activity), the decline in muscle mass and function represents the most dramatic and significant process of aging, making it a crucial and unavoidable issue in aging intervention. Sarcopenia is an age-related muscle disorder characterized by a decline in skeletal muscle mass, strength, or function associated with aging. The pathology of sarcopenia is complex, with various lifestyle factors contributing to its development, including malnutrition and lack of exercise. Furthermore, muscle is a high-energy-demand tissue, and mitochondrial dysfunction associated with aging, malnutrition, and insufficient physical activity is considered a major pathological feature.

[0009] Pompe disease (PD) is a rare inherited metabolic disorder caused by mutations in the acid α-glucosidase (GAA) gene, and it is inherited in an autosomal recessive manner. GAA gene mutations lead to a deficiency or reduction in GAA enzyme activity, preventing glycogen degradation and causing it to accumulate in the lysosomes of muscle cells. This damages the myocardium, skeletal muscle, and central nervous system, ultimately leading to heart failure or respiratory failure and death. Studies have shown that mitochondrial dysfunction, including a significant reduction in the number of mitochondria, impaired respiratory function and ATP production, and elevated levels of reactive oxygen species produced by intracellular mitochondria, participates in the pathological process.

[0010] Urolithin A is a secondary metabolite of ellagitannins, a natural polyphenol compound, and possesses various biological activities, such as antioxidant, anti-inflammatory, estrogen / androgen regulation, and autophagy promotion. Recent studies have shown that urolithin A has activities related to improving mitochondrial function, which is of great significance for the prevention and treatment of mitochondrial diseases. However, urolithin A has extremely poor solubility and very low oral bioavailability, making it difficult to formulate into a drug. Summary of the Invention

[0011] The purpose of this application is to provide compounds for improving mitochondrial function, preventing and treating aging and mitochondrial-related diseases, as well as their preparation methods and applications. This application modifies the structure of urolithin A using a prodrug method, resulting in urolithin A derivatives with good solubility, overcoming the problem of low bioavailability of urolithin A and improving its drug-like properties. The characteristics of urolithin A in improving mitochondrial function can be used to prevent and / or treat diseases related to mitochondrial function.

[0012] To achieve the above-mentioned objectives, this application provides the following technical solution:

[0013] This application provides a urolithin A derivative or a pharmaceutically acceptable salt thereof, the urolithin A derivative having the structure shown in Formula 1:

[0014] In Formula 1, X1 and X2 are independently selected from -O-, -S-, -NR3- or carbonyl, and at least one of X1 and X2 is carbonyl;

[0015] R1 is selected from -H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, -C(=O)R4, -C(=O)OR5 or -C(=O)NR6R7.

[0016] It consists of 3- to 10-membered nitrogen-containing heterocycles;

[0017] n is an integer from 0 to 18;

[0018] R2 is independently selected from deuterium, halogen, amino, nitro, carbonyl, ester, carboxyl, sulfonic acid, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C1-C6 amino, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocyclic alkyl; or R2 together with the attached nitrogen-containing heterocycle forms a substituted or unsubstituted spirocyclic ring or a substituted or unsubstituted bridged ring;

[0019] R3, R4, R5, R6, and R7 are independently selected from -H, deuterium, substituted or unsubstituted C1-C6 chain alkyl, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocyclic alkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl; or R6 and R7 together with the attached nitrogen atom form a substituted or unsubstituted C3-C7 heterocyclic alkyl.

[0020] Preferably, X1 and X2 are -O- and carbonyl groups, respectively.

[0021] Preferably, R1 is selected from -H, -CH3, -C(=O)CH3, -C(=O)OC(CH3)3, -C(=O)N(CH3)2 or

[0022] Preferably, R2 is selected from -CH3 or -N(CH3)2, or R2 together with the attached nitrogen-containing heterocycle forms an unsubstituted spiro ring or an unsubstituted bridged ring.

[0023] Preferably, the Selected from

[0024] Preferably, the Selected from

[0025] Preferably, the urolithin A derivative is selected from any one of the following compounds:

[0026] Preferably, the pharmaceutically acceptable salts of the urolithin A derivative include hydrochloride, sulfate, phosphate, hydrobromide, nitrate, salicylate, benzoate, C1-C6 fatty carboxylates, C1-C6 alkyl sulfonates, benzene sulfonates, p-toluene sulfonate, or camphor sulfonate.

[0027] This application provides a method for preparing the urolithin A derivative described in the above technical solution, comprising the following steps:

[0028] (I) When R1 is -H, and the When the urolithin A derivative is a nitrogen-containing heterocycle that does not contain a -NH- group, the preparation method includes the following steps:

[0029] Compound a, compound b, a base reagent, and an organic solvent were mixed and subjected to a substitution reaction to obtain compound c.

[0030] Compound c was subjected to a hydroxyl deprotection reaction to obtain compound d, wherein compound d is a compound with R1 being -H and The urolithin A derivative is a nitrogen-containing heterocycle that does not contain a -NH- group;

[0031] The structural formulas of compounds a, b, and c are shown below in sequence:

[0032] Z is a hydroxyl protecting group, X1, X2, R2 and n are defined as in Equation 1;

[0033] (II) When R1 is -H, and the When the urolithin A derivative is a nitrogen-containing heterocycle containing a -NH- group, the preparation method includes the following steps:

[0034] Compound a, compound b', a base reagent and an organic solvent were mixed and subjected to a substitution reaction to obtain compound c';

[0035] Compound c' is subjected to a hydroxyl deprotection reaction and an imine deprotection reaction to yield compound e, wherein compound e is R1-H and The urolithin A derivative is a nitrogen-containing heterocycle containing a -NH- group;

[0036] The difference between compound b' and compound b is that compound b' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle; the difference between compound c' and compound c is that compound c' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle.

[0037] (III) When R1 is not -H, the preparation method of the urolithin A derivative includes the following steps:

[0038] Compound d was prepared according to (I) or compound e was prepared according to (II);

[0039] In the presence of an alkaline reagent and an organic solvent, compound d or compound e is subjected to a substitution reaction with compound f to obtain the urolithin A derivative in which R1 is not -H;

[0040] The compound f is R4C(=O)X3, R5OC(=O)X3, R6R7NC(=O)X3 or R8X3, where X3 is a halogen atom and R8 is a substituted or unsubstituted C1-C6 chain alkyl, a substituted or unsubstituted C3-C7 cycloalkyl, a substituted or unsubstituted C3-C7 heterocycloalkyl, a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl.

[0041] This application provides the use of the urolithin A derivative or a pharmaceutically acceptable salt thereof described in the above-described technical solution in the preparation of medicaments for the prevention and / or treatment of diseases related to mitochondrial function.

[0042] Preferably, the diseases related to mitochondrial function include neurological diseases, aging, muscle diseases, or metabolic diseases.

[0043] Preferably, the neurodegenerative diseases include Friedreich's ataxia, amyotrophic lateral sclerosis (ALS), adrenoleukodystrophy, Alexander disease, Alpert disease, familial fatal insomnia, Huntington's disease, Kennedy's disease, Crabbe's disease, Lyme disease, Machado-Joseph disease, multiple sclerosis, multiple system atrophy, Niemann-Pick disease, Pick's disease, primary lateral sclerosis, progressive supranuclear palsy, Rafe-Syme disease, Sandhoff's disease, diffuse demyelinating sclerosis, spinocerebellar ataxia, subacute mixed spinal cord degeneration, toxic encephalopathy, disseminated spongiform encephalopathy, or ataxia-telangiectasia.

[0044] Preferably, the muscle disease includes sarcopenia or Duchenne muscular dystrophy.

[0045] Preferably, the metabolic disease includes obesity, hyperlipidemia, or Pompe disease.

[0046] This application provides a pharmaceutical composition comprising at least one of the urolithin A derivative and its pharmaceutically acceptable salt as described in the above technical solution, as well as a pharmaceutically acceptable excipient.

[0047] Preferably, the content of urolithiasis A derivative and / or its pharmaceutically acceptable salt in the pharmaceutical composition is 0.1 to 99.9 wt%.

[0048] This application provides a method for preventing and / or treating diseases related to mitochondrial function, comprising administering to a person preventing or treating diseases related to mitochondrial function an effective amount of at least one of the urolithin A derivative and its pharmaceutically acceptable salt, or the pharmaceutical composition described in the above-described technical solution.

[0049] This application provides a urolithin A derivative or a pharmaceutically acceptable salt thereof. This application modifies the structure of urolithin A via a prodrug approach, resulting in a urolithin A derivative with good solubility, overcoming the problem of low bioavailability of urolithin A and improving its drug-like properties. This allows for the prevention and / or treatment of diseases related to mitochondrial function by utilizing the urolithin A's ability to improve mitochondrial function. Attached Figure Description

[0050] Figure 1 shows the effect of CP-01 on the body weight of FA model mice;

[0051] Figure 2 shows the effect of CP-01 on the gripping force of the FA model mouse.

[0052] Figure 3 shows the effect of CP-01 on the FA model mouse rotarod test results;

[0053] Figure 4 shows the effect of CP-01 on the open field test of the FA model mouse.

[0054] Figure 5 shows the effect of CP-01 on behavioral tests in ALS model mice.

[0055] Figure 6 shows the effect of CP-01 on behavioral tests of MJD model mice;

[0056] Figure 7 shows the effect of CP-10 on behavioral tests in DMD model mice.

[0057] Figure 8 shows the effect of CP-10 on behavioral tests in ALS model mice.

[0058] Figure 9 shows the effect of CP-18 on behavioral tests in ALS model mice.

[0059] Figure 10 shows representative footprints from the gait test of CP-18 on ALS model mice.

[0060] Figure 11 shows the effect of CP-18 on the limb coordination ability of ALS model mice;

[0061] Figure 12 shows the effect of CP-18 on muscle damage in ALS model mice;

[0062] Figure 13 shows the effects of CP-18 on the level of inflammatory infiltration and fibrosis in the gastrocnemius muscle of ALS model mice;

[0063] Figure 14 shows the effect of CP-18 on the number of spinal cord neurons in ALS model mice (immunohistochemical diagram);

[0064] Figure 15 shows the effect of CP-18 on the number of spinal cord neurons in ALS model mice (statistical chart);

[0065] Figure 16 shows the effect of CP-18 on the level of spinal cord nerve inflammation in ALS model mice (immunohistochemical diagram);

[0066] Figure 17 shows the effect of CP-18 on the level of spinal cord nerve inflammation in ALS model mice (statistical chart);

[0067] Figure 18 shows the effects of CP-18 on the number and function of mitochondria in the gastrocnemius muscle of ALS model mice.

[0068] Figure 19 shows the effects of CP-18 on motor function and muscle strength in naturally aging mice.

[0069] Figure 20 shows representative footprints from the gait test of CP-18 in naturally aging mice.

[0070] Figure 21 shows the effect of CP-18 on limb coordination in naturally aging mice.

[0071] Figure 22 shows the effects of CP-18 on the inflammatory infiltration and fibrosis levels of the gastrocnemius muscle in naturally aged mice.

[0072] Figure 23 shows the effect of CP-18 on mitochondrial function in the gastrocnemius muscle of naturally aged mice.

[0073] Figure 24 shows the effects of CP-18 on mitochondrial copy number and telomere length in the gastrocnemius muscle of naturally aged mice.

[0074] Figure 25 shows the effect of CP-18 on the appearance of naturally aging mice.

[0075] Figure 26 shows the effect of CP-18 on the aging of the gastrocnemius muscle in naturally aging mice (representative image of the right hind limb of the mouse);

[0076] Figure 27 shows the effect of CP-18 on the aging of gastrocnemius muscle in naturally aging mice (statistical chart of gastrocnemius muscle wet weight ratio);

[0077] Figure 28 shows the effects of CP-18 on cognitive impairment in naturally aging mice.

[0078] Figure 29 shows a clustering diagram of differentially regulated proteins after CP-18 administration;

[0079] Figure 30 is a heatmap of cluster 1;

[0080] Figure 31 shows the clustering of differentially regulated proteins after CP-18 administration;

[0081] Figure 32 is a heatmap of cluster 2;

[0082] Figure 33 shows the effect of CP-18 on behavioral tests in PD model mice;

[0083] Figure 34 shows the effect of CP-18 on the expression of autophagy and lysosome-related proteins (lamp1 and LC3) in PD model mice (gastrocnemius muscle stained section);

[0084] Figure 35 shows the effect of CP-18 on the expression of autophagy and lysosome-related proteins (lamp1 and LC3) in PD model mice (myocardial stained sections). Detailed Implementation

[0085] This application provides a urolithin A derivative or a pharmaceutically acceptable salt thereof, the urolithin A derivative having the structure shown in Formula 1:

[0086] In Formula 1, X1 and X2 are independently selected from -O-, -S-, -NR3- or carbonyl, and at least one of X1 and X2 is carbonyl;

[0087] R1 is selected from -H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, -C(=O)R4, -C(=O)OR5 or -C(=O)NR6R7.

[0088] It consists of 3- to 10-membered nitrogen-containing heterocycles;

[0089] n is an integer from 0 to 18;

[0090] R2 is independently selected from deuterium, halogen, amino, nitro, carbonyl, ester, carboxyl, sulfonic acid, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C1-C6 amino, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocyclic alkyl; or R2 together with the attached nitrogen-containing heterocycle forms a substituted or unsubstituted spirocyclic ring or a substituted or unsubstituted bridged ring;

[0091] R3, R4, R5, R6, and R7 are independently selected from -H, deuterium, substituted or unsubstituted C1-C6 chain alkyl, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocyclic alkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl; or R6 and R7 together with the attached nitrogen atom form a substituted or unsubstituted C3-C7 heterocyclic alkyl.

[0092] In this application, X1 and X2 are independently selected from -O-, -S-, -NR3-, or carbonyl, and at least one of X1 and X2 is a carbonyl group. In this application, R3 is selected from -H, deuterium, substituted or unsubstituted C1-C6 alkyl groups, substituted or unsubstituted C3-C7 cycloalkyl groups, substituted or unsubstituted C3-C7 heterocycloalkyl groups, substituted or unsubstituted aryl groups, and substituted or unsubstituted heteroaryl groups. This application does not specifically limit the specific type of substituent in substituted C1-C6 alkyl groups, substituted C3-C7 cycloalkyl groups, substituted C3-C7 heterocycloalkyl groups, substituted aryl groups, and substituted heteroaryl groups. In this application, preferably, X1 and X2 are -O- and carbonyl groups, respectively; that is, when X1 is a carbonyl group, X2 is preferably -O-, and when X2 is a carbonyl group, X1 is preferably -O-.

[0093] In this application, R1 is selected from -H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, -C(=O)R4, -C(=O)OR5 or -C(=O)NR6R7. In this application, R4, R5, R6, and R7 are independently selected from -H, deuterium, substituted or unsubstituted C1-C6 alkyl chains, substituted or unsubstituted C3-C7 cycloalkyl chains, substituted or unsubstituted C3-C7 heterocyclic alkyl chains, substituted or unsubstituted aryl groups, substituted or unsubstituted heteroaryl groups, or R6 and R7 together with the attached nitrogen atom to form substituted or unsubstituted C3-C7 heterocyclic alkyl chains. This application does not specifically limit the specific types of substituents in substituted C1-C6 alkyl chains, substituted C3-C7 cycloalkyl chains, substituted C3-C7 heterocyclic alkyl chains, substituted aryl groups, and substituted heteroaryl groups. In this application, R4 is preferably -CH3; R5 is preferably -C(CH3)3; R6 is preferably -CH3; R7 is preferably -CH3; or R6 and R7 together with the attached nitrogen atom form a substituted 6-membered heterocyclic alkyl group, wherein the substituent is preferably -CH3. Specifically, the group formed by R6 and R7 together with the attached nitrogen atom is preferably... In this application, specifically, R1 is preferably -H, -CH3, -C(=O)CH3, -C(=O)OC(CH3)3, -C(=O)N(CH3)2 or

[0094] In this application, the stated It is a 3- to 10-membered nitrogen-containing heterocycle, specifically a 3-membered, 4-membered, 5-membered, 6-membered, 7-membered, 8-membered, 9-membered, or 10-membered nitrogen-containing heterocycle; specifically, the... Selected from

[0095] In this application, R2 is a substituent on the nitrogen-containing heterocycle, and n represents the number of R2 groups on the nitrogen-containing heterocycle.

[0096] In this application, n is an integer from 0 to 18, specifically 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17 or 18.

[0097] In this application, R2 is independently selected from deuterium, halogen, amino, nitro, carbonyl, ester, carboxyl, sulfonic acid, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C1-C6 amino, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocyclic alkyl, or R2 together with the attached nitrogen-containing heterocycle to form a substituted or unsubstituted spirocyclic ring or a substituted or unsubstituted bridged ring. This application does not specifically limit the specific types of substituents in the substituted C1-C6 alkyl, substituted C1-C6 amino, substituted C3-C7 cycloalkyl, substituted C3-C7 heterocyclic alkyl, substituted spirocyclic, and substituted bridged ring. Specifically, in this application, R2 is preferably -CH3 or -N(CH3)2, or R2 together with the attached nitrogen-containing heterocycle to form an unsubstituted spirocyclic ring or an unsubstituted bridged ring. Specifically, in this application, the... Preferred

[0098] In this application, the urolithin A derivative is specifically selected from any one of the following compounds:

[0099] In this application, the pharmaceutically acceptable salts of the urolithin A derivative preferably include hydrochloride, sulfate, phosphate, hydrobromide, nitrate, salicylate, benzoate, C1-C6 fatty carboxylates, C1-C6 alkyl sulfonates, benzene sulfonates, p-toluene sulfonate, or camphor sulfonate; the C1-C6 fatty carboxylates of this application preferably include oxalate, maleate, fumarate, citrate, succinate, or tartrate.

[0100] This application provides a method for preparing the urolithin A derivative described in the above technical solution. The preparation method is specifically selected based on the specific structure of the urolithin A derivative, and will be described in detail below. Unless otherwise specified, all raw materials used in this application are commercially available products well-known to those skilled in the art or prepared using methods well-known to those skilled in the art.

[0101] In this application, specifically, (I) when R1 is -H, and the When the urolithin A derivative is a nitrogen-containing heterocycle that does not contain a -NH- group, the preparation method includes the following steps:

[0102] Compound a, compound b, a base reagent, and an organic solvent were mixed and subjected to a substitution reaction to obtain compound c.

[0103] Compound c was subjected to a hydroxyl deprotection reaction to obtain compound d, wherein compound d is a compound with R1 being -H and The urolithin A derivative is a nitrogen-containing heterocycle that does not contain a -NH- group;

[0104] The structural formulas of compounds a, b, and c are shown below in sequence:

[0105] Z is a hydroxyl protecting group, X1, X2, R2 and n are defined as in Equation 1.

[0106] This application involves mixing compound a, compound b, a base reagent, and an organic solvent to conduct a substitution reaction to obtain compound c. In this application, Z in compound a is a hydroxyl protecting group, preferably including a benzyl protecting group (-Bn) or an oxycarbonyl protecting group (-Boc). In this application, the molar ratio of compound a (the preparation method of some compounds is detailed below) and compound b (the preparation method is detailed below) is preferably 1:0.5–2, more preferably 1:1–1.5. In this application, the base reagent is preferably a carbonate, more preferably potassium carbonate; the organic solvent is preferably acetonitrile; the molar ratio of compound a, base reagent, and organic solvent is preferably 1 mmol:1.5–2.5 mmol:15–22 mL, more preferably 1 mmol:2 mmol:20 mL. In this application, the temperature of the substitution reaction is preferably 75–85°C, more preferably 80°C; the time is preferably 5–7 h, more preferably 6 h. Following the substitution reaction, this application preferably extracts the resulting product system with ethyl acetate, dries the organic phase to anhydrous sodium sulfate, filters it, concentrates the filtrate, and then separates it by silica gel column chromatography to obtain compound c. In this application, the eluent used for silica gel column chromatography is preferably dichloromethane and methanol. The volume ratio of dichloromethane to methanol is selected according to actual needs, preferably 20–100:1, more preferably 50–80:1.

[0107] After obtaining compound c, this application subjects compound c to a hydroxyl deprotection reaction to obtain compound d, wherein compound d is the urolithin A derivative with R1 being H. This application preferably selects the appropriate hydroxyl deprotection reaction according to the specific type of the hydroxyl protecting group. Specifically, when the hydroxyl protecting group is -Bn, this application preferably removes it through a hydrogen reduction deprotection reaction; when the hydroxyl protecting group is -Boc, this application preferably removes it through an acid deprotection reaction. In this application, the hydrogen reduction deprotection reaction is preferably carried out in the presence of a hydrogenation catalyst, an organic solvent, and hydrogen. The hydrogenation catalyst is preferably palladium on carbon, and the amount used is not particularly required, as long as it can ensure the smooth progress of the reaction. The organic solvent is preferably a mixed solution of tetrahydrofuran and methanol, and the volume ratio of tetrahydrofuran to methanol is preferably 1:0.5 to 1.5, more preferably 1:1. The volume ratio of compound c to organic solvent is preferably 0.5 mmol:15 to 25 mL, more preferably 0.5 mmol:20 mL. The temperature of the hydrogen reduction deprotection reaction is preferably room temperature, and the time is preferably 10 to 15 h, more preferably 12 h. After the hydrogen reduction deprotection reaction, the obtained product system is preferably filtered through diatomaceous earth, and the filtrate is concentrated to obtain the urolithin A derivative. In this application, the acid deprotection reaction is preferably carried out in the presence of an ethyl hydrochloride solution and an organic solvent; the concentration of hydrochloric acid in the ethyl hydrochloride solution is preferably 3-5 mol / L, more preferably 4 mol / L; the organic solvent is preferably methanol; the ratio of compound c, ethyl hydrochloride solution and organic solvent is preferably 0.2-0.5 mmol: 1-1.5 mL: 5-15 mL; the temperature of the acid deprotection reaction is preferably room temperature, and the time is preferably 1.5-2.5 h, more preferably 2 h; after the acid deprotection reaction, this application preferably concentrates the obtained product system, dilutes the concentrate with ethyl acetate, washes it with saturated sodium bicarbonate solution, dries it with anhydrous sodium sulfate, filters it, concentrates the filtrate and separates it by silica gel column chromatography to obtain the urolithin A derivative. In this application, the eluent used for silica gel column chromatography is preferably dichloromethane and methanol. The volume ratio of dichloromethane to methanol is selected according to actual needs. The volume ratio of dichloromethane to methanol is preferably 20 to 100:1, and more preferably 30 to 50:1.

[0108] In this application, specifically, (II) when R1 is -H, and the When the urolithin A derivative is a nitrogen-containing heterocycle containing a -NH- group, the preparation method includes the following steps:

[0109] Compound a, compound b', a base reagent and an organic solvent were mixed and subjected to a substitution reaction to obtain compound c';

[0110] The compound c' was subjected to a hydroxyl deprotection reaction and an imino deprotection reaction to obtain compound e, wherein compound e is the urolithin A derivative with R1 being -H;

[0111] The difference between compound b' and compound b is that compound b' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle; the difference between compound c' and compound c is that compound c' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle.

[0112] This application involves mixing compound a, compound b', a base reagent, and an organic solvent to conduct a substitution reaction, yielding compound c'. In this application, compound b' differs from compound b in that compound b' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle; similarly, compound c' differs from compound c in that compound c' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle. The imino protecting group is preferably a tert-butoxycarbonyl protecting group (-Boc), a benzyloxycarbonyl protecting group (-Cbz), a benzyl protecting group (-Bn), a p-methoxybenzyl protecting group (-PMB), or a 2,4-dimethoxybenzyl protecting group (-DMB), more preferably -Boc. In this application, the ratio of compound a to compound b' (the preparation method is detailed below), the specific types and amounts of the base reagent and organic solvent, the substitution reaction conditions, and the post-treatment methods are preferably consistent with the substitution reaction using compound b as a starting material described above, and will not be repeated here.

[0113] After obtaining compound c', this application subjects compound c' to a hydroxyl deprotection reaction and an imino deprotection reaction to obtain compound e, wherein compound e is the urolithin A derivative with R1 being -H. In this application, when the hydroxyl protecting group is -Bn and the imino protecting group is -Boc, it is preferable to perform the hydroxyl deprotection reaction and the imino deprotection reaction sequentially, and more preferably to perform the hydrogen reduction deprotection reaction and the acid deprotection reaction sequentially. The specific reagents, reaction conditions, and post-treatment methods used are preferably consistent with the above-mentioned technical solutions and will not be repeated here. When both the hydroxyl protecting group and the imino protecting group are -Boc, it is preferable to perform the hydroxyl deprotection reaction and the imino deprotection reaction simultaneously, and more preferably to perform the acid deprotection reaction, that is, to simultaneously remove two -Boc groups through a single acid deprotection reaction. The specific reagents, reaction conditions, and post-treatment methods used are preferably consistent with the above-mentioned technical solutions and will not be repeated here.

[0114] In this application, specifically, (III) when R1 is not -H, the method for preparing the urolithin A derivative includes the following steps:

[0115] Compound d was prepared according to (I) or compound e was prepared according to (II);

[0116] In the presence of an alkaline reagent and an organic solvent, compound d or compound e is subjected to a substitution reaction with compound f to obtain the urolithin A derivative in which R1 is not -H;

[0117] The compound f is R4C(=O)X3, R5OC(=O)X3, R6R7NC(=O)X3 or R8X3, where X3 is a halogen atom and R8 is a substituted or unsubstituted C1-C6 chain alkyl, a substituted or unsubstituted C3-C7 cycloalkyl, a substituted or unsubstituted C3-C7 heterocycloalkyl, a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl.

[0118] After preparing compound d according to (I) or compound e according to (II), this application, in the presence of an alkaline reagent and an organic solvent, reacts compound d or compound e with compound f (X3 is a halogen atom, preferably -F, -Cl, -Br or -I) to obtain the urolithin A derivative whose R1 is not -H. In this application, the molar ratio of compound d (or compound e) to compound f is preferably 0.5:0.7-1.2, more preferably 0.5:0.8-1. In this application, the alkaline reagent is preferably a carbonate, more preferably potassium carbonate; the organic solvent is preferably acetonitrile; the ratio of the amount of compound d (or compound e), alkaline reagent, and organic solvent is preferably 0.5-1 mmol:0.8-2.5 mmol:15-25 mL, more preferably 0.5-1 mmol:1-2 mmol:20 mL. In this application, the temperature of the substitution reaction is preferably room temperature to reflux temperature; the time is preferably 5-7 h, more preferably 6 h. Following the substitution reaction, the resulting product system is preferably washed sequentially with dilute hydrochloric acid (preferably 1 mol / L) and saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, filtered, and the filtrate is concentrated and separated by silica gel column chromatography to obtain the urolithin A derivative. In this application, the eluent used for silica gel column chromatography is preferably dichloromethane and methanol, and the volume ratio of dichloromethane to methanol is selected according to actual needs. The preferred volume ratio is 20–100:1, more preferably 30–50:1.

[0119] In this application, it should be noted that when the hydroxyl protecting group in compound a is -Boc, and R1 in the target urolithin A derivative to be prepared is -OC(=O)OC(CH3)3, i.e. -OBoc, then compound c prepared by the substitution reaction of compound a and compound b is the urolithin A derivative with R1 -OBoc. It is not necessary to perform a hydroxyl deprotection reaction to prepare compound d and further use compound f for substitution reaction, as shown in Example 22 of this application for the preparation of compound Ua-f02.

[0120] The preparation methods of some compounds a, b, and b' in this application are described in detail below.

[0121] In this application, specifically, when X1 is a carbonyl group, X2 is -O-, and Z is -Bn, compound a has the structure shown in Formula 4; when X1 is -O-, X2 is a carbonyl group, and Z is -Boc, compound a has the structure shown in Formula 5.

[0122] In this application, the method for preparing the compound with the structure shown in Formula 4 preferably includes the following steps:

[0123] Compound I, benzyl bromide, a base reagent, and an organic solvent were mixed to carry out a substitution reaction, yielding compound II;

[0124] Compound II, a base reagent, water, and an organic solvent were mixed and subjected to an ester hydrolysis reaction to obtain compound III.

[0125] Compound III, resorcinol, a base reagent, copper sulfate, and water were mixed to carry out a ring-closure reaction, yielding the compound with the structure shown in Formula 4; the structural formulas of compounds I, II, and III are shown below in sequence:

[0126] This application involves mixing compound I, benzyl bromide, a base reagent, and an organic solvent to conduct a substitution reaction, yielding compound II. In this application, the molar ratio of compound I to benzyl bromide is preferably 10:18–25, more preferably 10:21. In this application, the base reagent is preferably a carbonate, more preferably potassium carbonate; the organic solvent is preferably N,N-dimethylformamide; the molar ratio of compound I, the base reagent, and the organic solvent is preferably 10 mmol:15–25 mmol:30–50 mL, more preferably 10 mmol:20 mmol:40 mL. In this application, the temperature of the substitution reaction is preferably 50–60 °C, more preferably 55 °C; the time is preferably 10–15 h, more preferably 12 h. After the substitution reaction, this application preferably extracts the resulting product system with a mixture of ethyl acetate and saturated ammonium chloride solution (the volume ratio of EA to saturated NH4Cl solution is preferably 2:1). The organic phase is dried over anhydrous sodium sulfate, filtered, and the filtrate is concentrated and separated by silica gel column chromatography to obtain compound II. In this application, the eluent used for silica gel column chromatography is preferably petroleum ether and ethyl acetate, and the volume ratio of petroleum ether to ethyl acetate is selected according to actual needs. The volume ratio of petroleum ether to ethyl acetate is preferably 10:1.

[0127] After obtaining compound II, this application mixes compound II, a base reagent, water, and an organic solvent to perform an ester hydrolysis reaction to obtain compound III. In this application, the base reagent is preferably lithium hydroxide; the organic solvent is preferably tetrahydrofuran; the preferred ratio of compound II, the base reagent, water, and the organic solvent is 5 mmol: 25–35 mmol: 8–12 mL: 15–25 mL, more preferably 5 mmol: 30 mmol: 10 mL: 20 mL. In this application, the preferred temperature for the ester hydrolysis reaction is -2 to 2°C, more preferably 0°C; the preferred time is 3.5–4.5 h, more preferably 4 h. After the ester hydrolysis reaction, this application preferably warms the resulting product system to room temperature, extracts with diethyl ether, adjusts the aqueous phase to pH = 1 with dilute hydrochloric acid (preferably 1 mol / L), extracts with ethyl acetate, dries the organic phase with anhydrous sodium sulfate, filters, and concentrates the filtrate to obtain compound III, which, without further purification, is directly used in the next reaction.

[0128] After obtaining compound III, this application mixes compound III, resorcinol, an alkaline reagent, copper sulfate, and water to carry out a ring-closure reaction to obtain the compound with the structure shown in Formula 4. In this application, the molar ratio of compound III to resorcinol is preferably 4:7-9, more preferably 4:8. In this application, the alkaline reagent is preferably sodium hydroxide; the preferred ratio of compound III, alkaline reagent, and water is 4 mmol:15-25 mmol:35-55 mL, more preferably 4 mmol:19.92 mmol:45 mL; in this application, copper sulfate acts as a catalyst, and its amount is sufficient to ensure the smooth progress of the reaction. In this application, the alkaline reagent and copper sulfate are preferably dissolved in water to obtain an alkaline reagent solution and a copper sulfate solution, respectively. Compound III and resorcinol are added sequentially to the alkaline reagent solution, and the mixture is refluxed and stirred for 25-35 min (preferably 30 min). Then, an aqueous solution of copper sulfate (preferably 5 wt%) is added, and the mixture is refluxed for another 10-15 h (preferably 12 h). After the ring-closing reaction, the resulting product system is preferably filtered, the filter cake is washed with dilute hydrochloric acid (preferably 1 mol / L), and vacuum dried to obtain the compound with the structure shown in Formula 4.

[0129] In this application, the method for preparing the compound with the structure shown in Formula 5 preferably includes the following steps:

[0130] The compound with the structure shown in Formula 4, ditert-butyl dicarbonate, imidazole, a base reagent and an organic solvent were mixed and subjected to a hydroxyl Boc protection reaction to obtain compound VII.

[0131] Compound VII was subjected to hydrogen reduction and debenzylation to obtain the compound with the structure shown in Formula 5.

[0132] This application involves mixing the compound with the structure shown in Formula 4, di-tert-butyl dicarbonate, imidazole, a base reagent, and an organic solvent to perform a hydroxyl Boc protection reaction to obtain compound VII. In this application, the molar ratio of the compound with the structure shown in Formula 4 to di-tert-butyl dicarbonate is preferably 2:3.5–4.5, more preferably 2:4. In this application, the base reagent is preferably an amine compound, more preferably triethylamine; the organic solvent is preferably dichloromethane; the molar ratio of the compound with the structure shown in Formula 4, imidazole, base reagent, and organic solvent is preferably 2 mmol:2.5–3.5 mmol:3.5–4.5 mmol:15–25 mL, more preferably 2 mmol:3 mmol:4 mmol:20 mL. In this application, the temperature of the hydroxyl Boc protection reaction is preferably room temperature, and the time is preferably 10–15 h, more preferably 12 h. After the hydroxyl Boc protection reaction, this application preferably extracts the resulting product system with ethyl acetate, dries the organic phase to anhydrous sodium sulfate, filters, concentrates the filtrate, and separates it by silica gel column chromatography to obtain compound VII. In this application, the eluent used for silica gel column chromatography is preferably petroleum ether and ethyl acetate, and the volume ratio of petroleum ether to ethyl acetate is selected according to actual needs, preferably 5:1.

[0133] After obtaining compound VII, this application subjectes compound VII to a hydrogen reduction debenzylation reaction to obtain the compound with the structure shown in Formula 5. In this application, the hydrogen reduction debenzylation reaction is preferably carried out in the presence of a hydrogenation catalyst, an organic solvent, and hydrogen. The hydrogenation catalyst is preferably palladium on carbon, and this application has no special requirements for its amount, as long as it can ensure the smooth progress of the reaction. The organic solvent is preferably a mixed solution of tetrahydrofuran and methanol, and the volume ratio of tetrahydrofuran to methanol is preferably 1:0.5 to 1.5, more preferably 1:1. The volume ratio of compound VII to organic solvent is preferably 1.5 mmol:15 to 25 mL, more preferably 1.5 mmol:20 mL. The temperature of the hydrogen reduction deprotection reaction is preferably room temperature, and the time is preferably 10 to 15 h, more preferably 12 h. After the hydrogen reduction deprotection reaction, this application preferably filters the obtained product system through diatomaceous earth and concentrates the filtrate to obtain the compound with the structure shown in Formula 5.

[0134] In this application, the method for preparing compound b preferably includes the following steps:

[0135] A nitrogen heterocyclic compound, an acyl chloride reagent, pyridine, and an organic solvent were mixed and subjected to an acyl chloride reaction to obtain compound b.

[0136] The structural formula of the nitrogen heterocyclic compound is shown below:

[0137] In this application, the acyl chloride reagent is preferably triphosgene; the organic solvent is preferably dichloromethane; the preferred ratio of the nitrogen heterocyclic compound, acyl chloride reagent, pyridine, and organic solvent is 2 mmol: 1.8–2.2 mmol: 2.5–3.5 mmol: 15–25 mL, more preferably 2 mmol: 2 mmol: 3 mmol: 20 mL. In this application, the preferred temperature for the acyl chloride reaction is room temperature, and the preferred time is 50–70 min, more preferably 60 min. After the acyl chloride reaction, the resulting product system is preferably washed with dilute hydrochloric acid (preferably 1 mol / L), the organic phase is dried over anhydrous sodium sulfate, filtered, and the filtrate is concentrated to obtain compound b, which can be directly used in the next reaction without further purification.

[0138] In this application, the preparation method of compound b' is preferably the same as that of compound b, the only difference being that the nitrogen-containing heterocyclic compound used to prepare compound b' contains an imino protecting group at the -NH- group of the nitrogen-containing heterocycle. Specifically, in this application, compound b' is used as... For example, the preferred preparation method is as follows:

[0139] N-Boc-piperazine, an acyl chloride reagent, pyridine, and an organic solvent were mixed and subjected to an acyl chloride reaction to obtain compound b'.

[0140] In this application, the specific types of acyl chloride reagent and organic solvent, the specific amounts of N-Boc-piperazine, acyl chloride reagent, pyridine and organic solvent, as well as the acyl chloride reaction conditions and post-treatment methods are preferably consistent with the above technical solutions, and will not be repeated here.

[0141] This application also preferably provides a method for preparing a pharmaceutically acceptable salt of the urolithin A derivative. Taking urolithin A derivative hydrochloride as an example, the method for preparing the urolithin A derivative hydrochloride preferably includes the following steps:

[0142] The urolithin A derivative, ethyl acetate hydrochloride solution, and organic solvent were mixed to carry out a salt formation reaction to obtain the urolithin A derivative hydrochloride.

[0143] In this application, the concentration of hydrochloric acid in the ethyl hydrochloric acid solution is preferably 3-5 mol / L, more preferably 4 mol / L; the organic solvent is preferably methanol; the ratio of the amount of urolithin A derivative, ethyl hydrochloric acid solution, and organic solvent is preferably 0.2 mmol:0.5-1.5 mL:5-15 mL, more preferably 0.2 mmol:1 mL:10 mL; the temperature of the salt-forming reaction is preferably room temperature, and the time is preferably 4-6 h, more preferably 5 h. After the salt-forming reaction, this application preferably filters the obtained product system, collects the filter cake, and obtains the urolithin A derivative hydrochloride.

[0144] This application provides the use of the urolithin A derivative or its pharmaceutically acceptable salt described in the above-described technical solution in the preparation of medicaments for the prevention and / or treatment of diseases related to mitochondrial function. In this application, the diseases related to mitochondrial function preferably include aging, nervous system diseases, muscle diseases, or metabolic diseases. In this application, the neurological diseases preferably include Friedreich's ataxia, amyotrophic lateral sclerosis (ALS), adrenoleukodystrophy, Alexander disease, Alper's disease, fatal familial insomnia, Huntington's disease, Kennedy's disease, Krabbe disease, Lyme disease, Machado-Joseph disease, multiple sclerosis, multiple system atrophy, Niemann-Pick disease, Pick's disease, primary lateral sclerosis, progressive supranuclear palsy, and Refsum disease. The diseases mentioned include: Sandhoff disease, diffuse myelinoclastic sclerosis, spinocerebellar ataxia, subacute combined degeneration of spinal cord, toxic encephalopathy, transmissible spongiform encephalopathy, or ataxia-telangiectasia; the muscle diseases preferably include sarcopenia or Duchenne muscular dystrophy; the metabolic diseases preferably include obesity, hyperlipidemia, or Pompe disease.

[0145] In this application, the drug specifically includes at least one of the urolithin A derivative and its pharmaceutically acceptable salt, as well as pharmaceutically acceptable excipients. This application does not specifically limit the type of pharmaceutically acceptable excipients; any excipient well known to those skilled in the art can be used. In this application, the content of the functional component (the urolithin A derivative and / or its pharmaceutically acceptable salt) in the drug is preferably 0.1–99.9 wt%, more preferably 1–99 wt%, and even more preferably 50–80 wt%. This application does not specifically limit the dosage form of the drug; any dosage form well known to those skilled in the art can be used.

[0146] This application provides a pharmaceutical composition comprising at least one of the urolithin A derivative and its pharmaceutically acceptable salt as described in the above-described technical solution, as well as a pharmaceutically acceptable excipient. In this application, the types of pharmaceutically acceptable excipients, the content of functional ingredients, and the range of applicable dosage forms are preferably consistent with the above-described solutions, and will not be repeated here.

[0147] The technical solutions of this application will be clearly and completely described below with reference to the embodiments therein. Obviously, the described embodiments are only a part of the embodiments of this application, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of this application without creative effort are within the scope of protection of this application.

[0148] Example 1: Synthesis of compound Ua-a01

[0149] Compound I (2.15 g, 10 mmol) was dissolved in N,N-dimethylformamide (DMF, 40 mL), followed by the sequential addition of potassium carbonate (K₂CO₃, 2.76 g, 20 mmol) and benzyl bromide (BnBr, 2.49 mL, 21 mmol). The mixture was stirred at 55 °C for 12 h. After the reaction was complete, the mixture was extracted with a mixture of ethyl acetate and saturated ammonium chloride solution (EA to saturated NH₄Cl solution, volume ratio 2:1). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (eluting PE:EA = 10:1) to give compound II (2.49 g, 63%). 1 H NMR (400MHz, CDCl3) δ7.51(d,J=8.8Hz,1H),7.49-7.29(m,11H),6.93(dd,J=8.8,3.1Hz,1H),5.35(s,2H),5.04(s,2H).

[0150] Compound II (1.98 g, 5 mmol) was dissolved in a mixed solution of tetrahydrofuran and methanol (THF to MeOH volume ratio 2:1, 30 mL). An aqueous solution of lithium hydroxide (0.72 g, 30 mmol) (10 mL) was added at 0 °C, and stirring was continued for 4 h. After the reaction was complete, the mixture was brought to room temperature and extracted twice with diethyl ether (Et₂O). The aqueous phase was adjusted to pH 1 with dilute hydrochloric acid (1 mol / L) and then extracted twice with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain the crude product, compound III. No further purification was required, and it was used directly in the next reaction.

[0151] Sodium hydroxide (0.8 g, 19.92 mmol) was dissolved in water (40 mL), and compound III (1.22 g, 4 mmol) and resorcinol (0.88 g, 8 mmol) were added sequentially. The mixture was refluxed and stirred for 30 min. Then, an aqueous solution of copper sulfate (5 wt%, 5 mL) was added, and the mixture was refluxed for another 12 h. After the reaction was complete, the mixture was filtered, washed with dilute hydrochloric acid (1 mol / L), and dried under vacuum to obtain compound IV (0.58 g, 46%). 1 H NMR(400MHz, DMSO-d6)δ7.96(d,J=8.0Hz,1H),7.70(d,J=9.9Hz,1H),7.61-7.55(m,1H),7.49( d,J=7.3Hz,2H),7.45-7.27(m,4H),6.38(d,J=8.7Hz,1H),6.18(d,J=1.6Hz,1H),5.20(s,2H).

[0152] N-methylpiperazine (200 mg, 2 mmol) was dissolved in anhydrous dichloromethane (DCM, 20 mL), followed by the sequential addition of pyridine (Py, 0.24 mL, 3 mmol) and triphosgene (BTC, 592 mg, 2 mmol). The reaction was carried out at room temperature for 1 h. After the reaction was complete, the mixture was washed with dilute hydrochloric acid (1 mol / L), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain a crude product. This crude product was then dissolved directly in acetonitrile (CH3CN, 20 mL), followed by the sequential addition of compound IV (318 mg, 1 mmol) and potassium carbonate (276 mg, 2 mmol). The mixture was stirred at 80 °C for 6 h. After the reaction was complete, the mixture was extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 50:1 as the eluent) to obtain compound V-01 (404 mg, 91%). 1H NMR (400MHz, CDCl3) δ8.02(d,J=8.9Hz,1H),7.98(d,J=8.7Hz,1H),7.92(d,J=2.7Hz,1H),7.49(dd,J=8.4,3.0Hz,3H),7.4 4(t,J=7.3Hz,2H),7.38(t,J=7.1Hz,1H),7.20-7.12(m,2H),5.22(s,2H),3.71(m,4H),2.53(t,J=5.0Hz,4H),2.40(s,3H).

[0153] Compound V-01 (222 mg, 0.5 mmol) was dissolved in a mixed solution of tetrahydrofuran and methanol (THF to MeOH volume ratio 1:1, 20 mL), and palladium on carbon (Pd / C, Pd content 10 wt%, 2 g) was added. Air was replaced with nitrogen, followed by nitrogen replacement, and the reaction was carried out under a hydrogen atmosphere at room temperature with stirring for 12 h. After the reaction was complete, the mixture was filtered through diatomaceous earth, and the filtrate was concentrated to obtain compound Ua-a01 (83 mg, 47%). 1 H NMR (400MHz, DMSO-d6) δ10.42(s,1H),8.24(t,J=8.7Hz,2H),7.57(d,J=2.7Hz,1H),7.37(dd,J=8.7,2.7Hz, 1H),7.23(d,J=2.3Hz,1H),7.16(dd,J=8.7,2.4Hz,1H),3.60(s,2H),3.45(s,2H),2.38(s,4H),2.23(s,3H).

[0154] Example 2 Synthesis of compound Ua-a02

[0155] The preparation method is basically the same as that in Example 1, except that 4-dimethylaminopiperidine is used instead of N-methylpiperazine to obtain compound Ua-a02 (105 mg, 55%). 1 H NMR (400MHz, DMSO-d6) δ10.46(s,1H),8.25(t,J=8.7Hz,2H),7.57(d,J=2.7Hz,1H),7.36(dd,J=8.7,2.7Hz,1H),7. 23(d,J=2.4Hz,1H),7.15(dd,J=8.7,2.4Hz,1H),4.33(m,4H),2.93(m,1H),2.77(s,6H),2.11(s,2H),1.70(s,2H).

[0156] Example 3 Synthesis of compound Ua-a03

[0157] The preparation method is basically the same as that in Example 1, except that N-methylperpiperazine is used instead of N-methylpiperazine to obtain compound Ua-a03 (126 mg, 66%). 1 H NMR (400MHz, DMSO-d6) δ10.50(s,1H),8.23(dd,J=8.8,7.3Hz,2H),7.60(d,J=2.7Hz,1H),7.40(dd,J=8.8,2.7Hz,1H),7.22(d, J=2.4Hz,1H),7.15(dd,J=8.8,2.4Hz,1H),3.72-3.60(m,2H),3.58-3.46(m,2H),2.80-2.47(m,5H),2.34(s,2H),1.88(m,2H).

[0158] Example 4: Synthesis of compound Ua-a04

[0159] The preparation method is basically the same as that in Example 1, except that (R)-octahydropyrrolo[1,2-A]pyrazine is used instead of N-methylpiperazine to obtain compound Ua-a04 (80 mg, 42%). 1 H NMR (400MHz, DMSO-d6) δ10.44(s,1H),8.25(t,J=8.6Hz,2H),7.57(d,J=2.7Hz,1H),7.38(dd,J=8.8,2.7Hz,1H),7.24(d,J=2.4Hz,1H),7.17(dd,J=8 .7,2.4Hz,1H),4.30-3.96(m,2H),3.26-2.92(m,3H),2.84-2.60(m,1H),2 .19-2.05(m,2H),2.02-1.89(m,1H),1.87-1.59(m,3H),1.38-1.26(m,1H).

[0160] Example 5: Synthesis of compound Ua-b01

[0161] The synthesis steps of compound IV are as described in Example 1.

[0162] N-Boc-piperazine (372 mg, 2 mmol) was dissolved in anhydrous dichloromethane (DCM, 20 mL), followed by the sequential addition of pyridine (Py, 0.24 mL, 3 mmol) and triphosgene (BTC, 592 mg, 2 mmol). The reaction was carried out at room temperature for 1 h. After the reaction was complete, the mixture was washed with dilute hydrochloric acid (1 mol / L), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain a crude product. This crude product was then dissolved directly in acetonitrile (CH3CN, 20 mL), followed by the sequential addition of compound IV (318 mg, 1 mmol) and potassium carbonate (276 mg, 2 mmol). The mixture was stirred at 80 °C for 6 h. After the reaction was complete, the mixture was extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 80:1 as the eluent) to obtain compound V-05 (498 mg, 94%).

[0163] Compound V-05 (265 mg, 0.5 mmol) was dissolved in a mixed solution of tetrahydrofuran and methanol (THF to MeOH volume ratio 1:1, 20 mL), and palladium on carbon (Pd / C, 10%, 2 g) was added. Air was replaced with nitrogen, followed by nitrogen replacement, and the reaction was carried out under a hydrogen atmosphere with stirring at room temperature for 12 h. After the reaction was complete, the mixture was filtered through diatomaceous earth, and the filtrate was concentrated to obtain compound VI-05 (97 mg, 44%).

[0164] Compound VI-05 (88 mg, 0.2 mmol) was dissolved in methanol (10 mL), and ethyl acetate hydrochloride solution (4 mol / L, 1 mL) was added. The mixture was stirred at room temperature for 2 h. After the reaction was completed, the solution was concentrated, diluted with ethyl acetate, washed with saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 20:1 as the eluent) to obtain compound Ua-b01 (61 mg, 89%). 1 H NMR (400MHz, DMSO-d6) δ10.73(s,1H),8.27(d,J=8.6Hz,2H),7.63(d,J=2.3Hz,1H),7.44(dd, J=8.4,2.3Hz,1H),7.30(s,1H),7.22(d,J=8.1Hz,1H),3.53(s,2H),3.38(s,2H),3.18(s,4H).

[0165] Example 6 Synthesis of compound Ua-b02

[0166] The preparation method is basically the same as that in Example 5, except that N-1-Boc-2-methylpiperazine is used instead of N-Boc-piperazine to obtain compound Ua-b02 (67 mg, 95%). 1H NMR (400MHz, DMSO-d6) δ10.74(s,1H),8.27(d,J=8.6Hz,2H),7.65(d,J=2.4Hz,1H),7.44(dd,J=8.4,2. 4Hz,1H),7.31(s,1H),7.22(d,J=8.2Hz,1H),4.49-4.35(m,1H),3.33-2.84(m,6H),1.33-1.25(m,3H).

[0167] Example 7 Synthesis of compound Ua-b03

[0168] The preparation method was basically the same as in Example 5, except that N-Boc-piperazine was replaced with tert-butyl 4,7-diazaspiro[2.5]octane-4-carboxylate to obtain compound Ua-b03 (63 mg, 86%). 1 H NMR (400MHz, DMSO-d6) δ10.73(s,1H),8.26(d,J=8.6Hz,2H),7.64(d,J=2.3Hz,1H),7.44(dd,J=8.4,2. 3Hz,1H),7.32(s,1H),7.22(d,J=8.1Hz,1H),3.63(m,2H),3.52(s,2H),2.85-2.78(m,2H),0.44(s,4H).

[0169] Example 8 Synthesis of compound Ua-b04

[0170] The preparation method is basically the same as that in Example 5, except that (1S,4S)-2-Boc-2,5-diazabicyclo[2.2.1]heptane is used to replace N-Boc-piperazine to obtain compound Ua-b04 (64 mg, 91%). 1 H NMR(400MHz,DMSO-d6)δ10.64(s,1H),8.16(ddd,J=8.7,6.7,1.6Hz,2H),7.50( d,J=2.7Hz,1H),7.30(dd,J=8.7,2.7Hz,1H),7.15(dd,J=8.2,2.4Hz,1H),7.09 (ddd,J=8.6,6.2,2.4Hz,1H),4.49(s,1H),4.30(s,1H),3.60(d,J=9.7Hz,1H), 3.50-3.14(m,2H),3.00-2.74(m,2H),1.83-1.66(m,1H),1.58(t,J=9.6Hz,1H).

[0171] Example 9: Synthesis of compound Ua-b05

[0172] The preparation method was basically the same as in Example 5, except that N-Boc-piperazine was replaced with cis-2-Boc-hexahydropyrrolo[3,4-c]pyrrole to obtain compound Ua-b05 (67 mg, 92%). 1 H NMR (400MHz, DMSO-d6) δ10.69(s,1H),8.24(d,J=8.7Hz,2H),7.59(d,J=2.3Hz,1H),7.41(dd,J=8.4,2.3Hz,1H),7.25(s,1H),7.1 8(d,J=8.1Hz,1H),4.05-3.88(m,2H),3.60-3.48(m,2H),2.90(s,2H),2.81(s,2H),2.68(d,J=3.1Hz,1H),2.65(d,J=3.1Hz,1H).

[0173] Example 10 Synthesis of compound Ua-c01

[0174] Compound Ua-a01 (177 mg, 0.5 mmol) was dissolved in acetonitrile (20 mL), and potassium carbonate (138 mg, 1 mmol) and dimethylcarbamoyl chloride (0.07 mL, 0.8 mmol) were added sequentially. The mixture was stirred under reflux for 6 h. After the reaction was completed, the mixture was washed successively with dilute hydrochloric acid (1 mol / L) and saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 40:1 as the eluent) to obtain compound Ua-c01 (176 mg, 83%). 1 H NMR (400MHz, CDCl3) δ8.09(d,J=2.6Hz,1H),8.05(d,J=8.8Hz,1H),7.99(d,J=8.5Hz,1H),7.64(dd,J=8.8,2.6Hz,1H),7.1 8-7.12(m,2H),3.72(t,J=5.2Hz,2H),3.62(t,J=5.0Hz,2H),3.14(s,3H),3.05(s,3H),2.49(t,J=5.0Hz,4H),2.36(s,3H).

[0175] Example 11 Synthesis of compound Ua-co2

[0176] The preparation method was basically the same as in Example 10, except that acetyl chloride was used instead of dimethylcarbamoyl chloride to obtain compound Ua-c02 (144 mg, 73%). 1 H NMR (400MHz, CDCl3) δ8.03-7.94(m,2H),7.89(d,J=8.5Hz,1H),7.48(dd,J=8.7,2.6Hz,1H),7.14- 6.97(m,2H),3.68(t,J=5.2Hz,2H),3.57(t,J=5.2Hz,2H),2.49(s,4H),2.30(s,3H),2.28(s,3H).

[0177] Example 12 Synthesis of compound Ua-co3

[0178] Compound Ua-a01 (177 mg, 0.5 mmol) was dissolved in acetonitrile (20 mL), and potassium carbonate (138 mg, 1 mmol) and iodomethane (0.05 mL, 0.8 mmol) were added sequentially. The mixture was refluxed and stirred for 6 h. After the reaction was completed, the mixture was washed with dilute hydrochloric acid (1 mol / L), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 50:1 as the eluent) to obtain compound Ua-c03 (162 mg, 88%). 1 H NMR(400MHz, CDCl3) δ7.94(dd,J=10.1,8.7Hz,2H),7.76(d,J=2.8Hz,1H),7.37(dd,J=8.8,2.8Hz,1H),7.18 -7.05(m,2H),3.93(s,3H),3.72(t,J=5.2Hz,2H),3.62(t,J=5.2Hz,2H),2.49(t,J=5.1Hz,2H),2.36(s,3H).

[0179] Example 13 Synthesis of compound Ua-d01

[0180] The synthesis steps of compound IV are as described in Example 1.

[0181] Compound IV (636 mg, 2 mmol) was dissolved in dichloromethane (20 mL), and imidazole (204 mg, 3 mmol), triethylamine (0.55 mL, 4 mmol), and di-tert-butyl dicarbonate (0.9 mL, 4 mmol) were added sequentially. The mixture was stirred at room temperature for 12 h. After the reaction was complete, the mixture was extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using PE:EA = 5:1 as the eluent) to give compound VII (744 mg, 89%).1 H NMR (400MHz, CDCl3) δ8.02(dd,J=13.0,8.8Hz,2H),7.94(d,J=2.7Hz,1H),7.50(dd,J=8.9,2.5Hz,3H),7.47- 7.42(m,2H),7.41-7.37(m,1H),7.26(d,J=2.3Hz,1H),7.21(dd,J=9.0,2.0Hz,1H),5.23(s,2H),1.61(s,9H).

[0182] Compound VII (627 mg, 1.5 mmol) was dissolved in a mixed solution of tetrahydrofuran and methanol (THF to MeOH volume ratio 1:1, 20 mL), and palladium on carbon (Pd / C, 10%, 2 g) was added. Air was purged with nitrogen, followed by nitrogen purging, and the reaction was carried out under a hydrogen atmosphere at room temperature with stirring for 12 h. After the reaction was complete, the mixture was filtered through diatomaceous earth, and the filtrate was concentrated to obtain crude compound VIII, which was used directly in the next reaction without further purification.

[0183] N-methylpiperazine (200 mg, 2 mmol) was dissolved in anhydrous dichloromethane (DCM, 20 mL), followed by the sequential addition of pyridine (Py, 0.24 mL, 3 mmol) and triphosgene (BTC, 592 mg, 2 mmol). The reaction was carried out at room temperature for 1 h. After the reaction was complete, the mixture was washed with dilute hydrochloric acid (1 mol / L), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain a crude product. This crude product was then dissolved directly in acetonitrile (CH3CN, 20 mL), followed by the sequential addition of compound VIII (328 mg, 1 mmol) and potassium carbonate (276 mg, 2 mmol). The mixture was stirred at 80 °C for 6 h. After the reaction was complete, the mixture was extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 50:1 as the eluent) to obtain compound IX-01 (426 mg, 94%). 1 H NMR (400MHz, DMSO-d6) δ8.46(d,J=8.9Hz,1H),8.40(d,J=8.9Hz,1H),7.96(d,J=2.5Hz,1H),7.77(dd,J=8.8,2.5 Hz,1H),7.40(d,J=2.3Hz,1H),7.30(dd,J=8.7,2.3Hz,1H),3.56(m,4H),2.41(s,4H),2.25(s,3H),1.53(s,9H).

[0184] Compound IX-01 (227 mg, 0.5 mmol) was dissolved in methanol (10 mL), and ethyl acetate hydrochloride solution (4 mol / L, 1.5 mL) was added. The mixture was stirred at room temperature for 2 h. After the reaction was complete, the solution was concentrated, diluted with ethyl acetate, washed with saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 30:1 as the eluent) to obtain compound Ua-d01 (120 mg, 68%).

[0185] 1 H NMR (400MHz, DMSO-d6) δ10.36(s,1H),8.32(d,J=8.9Hz,1H),8.16(d,J=8.9Hz,1H),7.97(d,J=2.6Hz,1H),7.71(dd,J=8 .8, 2.6Hz, 1H), 6.88 (dd, J=8.7, 2.4Hz, 1H), 6.79 (d, J=2.4Hz, 1H), 3.60 (s, 2H), 3.44 (s, 2H), 2.38 (s, 4H), 2.21 (s, 3H).

[0186] Example 14 Synthesis of compound Ua-d02

[0187] The preparation method was basically the same as in Example 13, except that 4-dimethylaminopiperidine was used instead of N-methylpiperazine to obtain compound Ua-d02 (120 mg, 63%).

[0188] 1 H NMR (400MHz, DMSO-d6) δ10.37(s,1H),8.32(d,J=8.9Hz,1H),8.16(d,J=8.9Hz,1H),7.97(d,J=2.6Hz,1H),7.72(dd,J=8.8,2. 6Hz,1H),6.88(dd,J=8.7,2.4Hz,1H),6.79(d,J=2.4Hz,1H),4.32(m,4H),2.93(m,1H),2.76(s,6H),2.11(s,2H),1.69(s,2H).

[0189] Example 15 Synthesis of compound Ua-d03

[0190] The preparation method was basically the same as in Example 13, except that N-methylperiperazine was used instead of N-methylpiperazine to obtain compound Ua-d03 (94 mg, 51%).

[0191] 1H NMR (400MHz, DMSO-d6) δ10.37(s,1H),8.32(d,J=8.8Hz,1H),8.15(d,J=8.8Hz,1H),7.97(d,J=2.6Hz,1H),7.72(dd,J=8.8,2.6Hz,1H), 6.86(dd,J=8.7,2.4Hz,1H),6.77(d,J=2.4Hz,1H),3.72-3.60(m,2H),3.58-3.46(m,2H),2.80-2.47(m,5H),2.34(s,2H),1.88(m,2H).

[0192] Example 16 Synthesis of compound Ua-d04

[0193] The preparation method was basically the same as in Example 13, except that (R)-octahydropyrrolo[1,2-A]pyrazine was used instead of N-methylpiperazine to obtain compound Ua-d04 (125 mg, 66%).

[0194] 1 H NMR (400MHz, DMSO-d6) δ10.38(s,1H),8.32(d,J=8.9Hz,1H),8.15(d,J=8.9Hz,1 H),7.97(d,J=2.6Hz,1H),7.72(dd,J=8.8,2.6Hz,1H),6.86(dd,J=8.7,2.4Hz,1 H),6.77(d,J=2.4Hz,1H),4.32-3.96(m,2H),3.26-2.92(m,3H),2.84-2.60(m,1 H),2.19-2.05(m,2H),2.02-1.89(m,1H),1.87-1.59(m,3H),1.38-1.25(m,1H).

[0195] Example 17 Synthesis of compound Ua-e01

[0196] The synthesis steps of compound VIII are described in Example 13.

[0197] N-Boc-piperazine (372 mg, 2 mmol) was dissolved in anhydrous dichloromethane (DCM, 20 mL), followed by the sequential addition of pyridine (Py, 0.24 mL, 3 mmol) and triphosgene (BTC, 592 mg, 2 mmol). The reaction was carried out at room temperature for 1 h. After the reaction was complete, the mixture was washed with dilute hydrochloric acid (1 mol / L), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain a crude product. This crude product was then dissolved directly in acetonitrile (CH3CN, 20 mL), followed by the sequential addition of compound VIII (328 mg, 1 mmol) and potassium carbonate (276 mg, 2 mmol). The mixture was stirred at 80 °C for 6 h. After the reaction was complete, the mixture was extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 100:1 as the eluent) to obtain compound IX-05 (491 mg, 91%).

[0198] Compound IX-05 (108 mg, 0.2 mmol) was dissolved in methanol (10 mL), and ethyl acetate hydrochloride solution (4 mol / L, 1 mL) was added. The mixture was stirred at room temperature for 2 h. After the reaction was completed, the mixture was concentrated, diluted with ethyl acetate, and washed with saturated sodium bicarbonate solution. The washed product was then concentrated and separated by silica gel column chromatography (using DCM:MeOH = 20:1 as the eluent) to obtain compound Ua-e01 (60 mg, 88%).

[0199] 1 H NMR (400MHz, DMSO-d6) δ10.38(s,1H),8.32(d,J=8.9Hz,1H),8.15(d,J=8.9Hz,1H),7.97(d,J=2.6Hz,1H),7.72( dd,J=8.8,2.6Hz,1H),6.86(dd,J=8.7,2.4Hz,1H),6.77(d,J=2.4Hz,1H),3.54(s,2H),3.38(s,2H),3.17(s,4H).

[0200] Example 18 Synthesis of compound Ua-e02

[0201] The preparation method is basically the same as that in Example 17, except that N-1-Boc-2-methylpiperazine is used instead of N-Boc-piperazine to obtain compound Ua-e02 (78 mg, 44%).

[0202] 1H NMR (400MHz, DMSO-d6) δ10.31(s,1H),8.32(d,J=8.9Hz,1H),8.15(d,J=8.9Hz,1H),7.96(d,J=2.6Hz,1H),7.72(dd,J=8.8 ,2.6Hz,1H),6.87(dd,J=8.7,2.4Hz,1H),6.74(d,J=2.4Hz,1H),4.51-4.36(m,1H),3.31-2.82(m,6H),1.33-1.23(m,3H).

[0203] Example 19 Synthesis of compound Ua-e03

[0204] The preparation method was basically the same as in Example 17, except that N-Boc-piperazine was replaced with tert-butyl 4,7-diazaspiro[2.5]octane-4-carboxylate to obtain compound Ua-e03 (108 mg, 59%).

[0205] 1 H NMR (400MHz, DMSO-d6) δ10.35(s,1H),8.29(d,J=8.8Hz,1H),8.13(d,J=8.8Hz,1H),7.94(d,J=2.6Hz,1H),7.71(dd,J=8.8 ,2.6Hz,1H),6.86(dd,J=8.7,2.4Hz,1H),6.74(d,J=2.4Hz,1H),3.61(m,2H),3.52(s,2H),2.85-2.78(m,2H),0.45(s,4H).

[0206] Example 20 Synthesis of compound Ua-e04

[0207] The preparation method is basically the same as that in Example 17, except that (1S,4S)-2-BOC-2,5-diazabicyclo[2.2.1]heptane is used to replace N-Boc-piperazine to obtain compound Ua-e04 (83mg, 47%).

[0208] 1H NMR (400MHz, DMSO-d6) δ10.37(s,1H),8.30(d,J=8.8Hz,1H),8.15(d,J=8.8Hz, 1H),7.95(d,J=2.6Hz,1H),7.71(dd,J=8.8,2.6Hz,1H),6.87(dd,J=8.7,2.4Hz ,1H),6.74(d,J=2.4Hz,1H),4.50(s,1H),4.30(s,1H),3.61(d,J=9.7Hz,1H),3 .52-3.16(m,2H),3.00-2.74(m,2H),1.85-1.66(m,1H),1.58(t,J=9.6Hz,1H).

[0209] Example 21 Synthesis of compound Ua-f01

[0210] N-methylpiperazine (0.22 mL, 2 mmol) was dissolved in anhydrous dichloromethane (DCM, 20 mL), followed by the sequential addition of pyridine (Py, 0.24 mL, 3 mmol) and triphosgene (BTC, 592 mg, 2 mmol). The reaction was carried out at room temperature for 1 h. After the reaction was complete, the mixture was washed with dilute hydrochloric acid (1 mol / L), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain a crude product. This crude product was then dissolved directly in acetonitrile (CH3CN, 20 mL), followed by the sequential addition of compound Ua-d01 (354 mg, 1 mmol) and potassium carbonate (276 mg, 2 mmol). The mixture was stirred at 80 °C for 6 h. After the reaction was complete, the mixture was extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 20:1 as the eluent) to obtain compound Ua-f01 (408 mg, 85%). 1 H NMR (400MHz, CDCl3) δ8.10(d,J=2.5Hz,1H),8.06(d,J=8.9Hz,1H),8.00(d,J=8.5Hz,1H),7.63(dd,J=8.7,2. 6Hz,1H),7.19-7.09(m,2H),3.73(d,J=5.8Hz,4H),3.62(t,J=5.1Hz,4H),2.49(t,J=5.0Hz,8H),2.36(s,6H).

[0211] Example 22 Synthesis of compound Ua-f02

[0212] The synthetic route of compound Ua-f02 is the same as that of intermediate compound IX-01 in Example 13 (426 mg, 94%). 1H NMR (400MHz, DMSO-d6) δ8.46(d,J=8.9Hz,1H),8.40(d,J=8.9Hz,1H),7.96(d,J=2.5Hz,1H),7.77(dd,J=8.8,2.5 Hz,1H),7.40(d,J=2.3Hz,1H),7.30(dd,J=8.7,2.3Hz,1H),3.56(s,4H),2.41(s,4H),2.25(s,3H),1.53(s,9H).

[0213] Example 23 Synthesis of compound Ua-f03

[0214] Compound Ua-d01 (177 mg, 0.5 mmol) was dissolved in acetonitrile (20 mL), and potassium carbonate (138 mg, 1 mmol) and acetyl chloride (0.07 mL, 0.8 mmol) were added sequentially. The mixture was stirred at room temperature for 6 h. After the reaction was completed, the mixture was washed successively with dilute hydrochloric acid (1 mol / L) and saturated sodium bicarbonate solution, then dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 30:1 as the eluent) to obtain compound Ua-f03 (174 mg, 88%).

[0215] 1 H NMR (400MHz, DMSO-d6) δ8.44(d,J=8.9Hz,1H),8.39(d,J=8.9Hz,1H),7.96(d,J=2.5Hz,1H),7.76(dd,J=8.8,2.5 Hz,1H),7.40(d,J=2.3Hz,1H),7.30(dd,J=8.7,2.3Hz,1H),3.56(s,4H),2.41(s,4H),2.32(s,3H),2.25(s,3H).

[0216] Example 24 Synthesis of compound Ua-f04

[0217] Compound Ua-d01 (177 mg, 0.5 mmol) was dissolved in acetonitrile (20 mL), followed by the addition of potassium carbonate (138 mg, 1 mmol) and iodomethane (0.05 mL, 0.8 mmol). The mixture was refluxed (80 °C) and stirred for 6 h. After the reaction was complete, the mixture was washed successively with dilute hydrochloric acid (1 mol / L) and saturated sodium bicarbonate solution, then dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and separated by silica gel column chromatography (using DCM:MeOH = 30:1 as the eluent) to obtain compound Ua-f04 (167 mg, 91%).

[0218] 1 H NMR (400MHz, DMSO-d6) δ8.44(d,J=8.9Hz,1H),8.39(d,J=8.9Hz,1H),7.96(d,J=2.5Hz,1H),7.76(dd,J=8.8,2.5Hz ,1H),7.40(d,J=2.3Hz,1H),7.30(dd,J=8.7,2.3Hz,1H),3.81(s,3H),3.61-3.47(m,4H),2.41(s,4H),2.25(s,3H).

[0219] Example 25 Synthesis of compound CP-01 (Ua-a01 hydrochloride)

[0220] Compound Ua-a01 (70 mg, 0.2 mmol) was dissolved in methanol (10 mL), and ethyl acetate hydrochloride solution (4 mol / L, 1 mL) was added. The mixture was stirred at room temperature for 5 h. After the reaction was completed, the mixture was filtered, and the filter cake was collected to give a white solid compound CP-01 (81 mg, 96%). 1 H NMR (400MHz, DMSO-d6) δ11.19(s,1H),10.54(s,1H),8.26(dd,J=8.9,1.7Hz,2H),7.59(d,J=2.7Hz,1H),7.40(dd,J=8.7,2.7Hz ,1H),7.28(d,J=2.3Hz,1H),7.21(dd,J=8.7,2.3Hz,1H),4.45-3.96(m,2H),3.53-3.41(m,4H),3.26-3.07(m,2H),2.81(s,3H).

[0221] Following the method of Example 25, the hydrochlorides of the compounds in Examples 2 to 24 were prepared.

[0222] Test Example 1: Water Solubility Test

[0223] To determine the water solubility of the compounds in this application, the following compounds were dissolved in water at room temperature, and their solubility was observed. The results are shown in Table 1. As can be seen from Table 1, the water solubility of the compounds provided in this application is significantly improved compared to urolithin A.

[0224] Table 1. Water solubility test results of the compounds in this application.

[0225] Test Example 2: Cell Viability Test

[0226] The cell viability assessment method in this test case is as follows:

[0227] Mitophagy: Mouse neuroblastoma-derived N2a cells were passaged in confocal microscopy dishes, the culture medium was discarded, and the cells were washed twice with HBBS. Mitophagy dye (100 nmol / L) was prepared in advance according to the manufacturer's instructions, and the cells were incubated at 37°C for 30 min in the dark. After staining, the cells were gently washed twice with HBBS, ensuring complete darkness throughout the process, and the cells were treated for 24 h as designed. After the treatment, the cells were washed three times with HBBS and treated in the dark. Lyso dye (1 μmol / L) was prepared in advance and incubated at 37°C for 30 min. After staining, the cells were gently washed twice with PBS, stained with Hoechst for 10 min, and the colocalization of mitochondrial autophagosomes was analyzed using a confocal fluorescence microscope.

[0228] Mitochondrial ATP content detection: Lysis buffer (200 μL) was added to N2a cells, and the cells were homogenized using an ultrasonic homogenizer. After lysis, a portion of the sample was boiled for 2 min to fully release ATP. The remaining sample was centrifuged for 20 min (4℃, 12000g), and the supernatant was collected for BCA protein quantification. Standard curve preparation: Concentration gradients of 0.01 μM, 0.03 μM, 0.1 μM, 0.3 μM, 1 μM, 3 μM, and 10 μM were set. ATP detection working solution (100 μL) was added to a 96-well black plate and incubated at room temperature for 3–5 min. Then, 20 μL of sample or standard was added to each well, and the mixture was quickly mixed with a pipette. The RLU value was measured using a chemiluminescence analyzer. ATP levels were calculated in nmol / mg protein.

[0229] The specific test results are shown in Table 2.

[0230] Table 2 Results of cell viability test

[0231] Test Example 3: Pharmacokinetic Study

[0232] This test case specifically selected three representative compounds with different modifications (CP-01: Ua-a01 hydrochloride; CP-10: Ua-d01 hydrochloride; CP-18: Ua-f01 hydrochloride) for pharmacokinetic studies. The specific steps are as follows:

[0233] 1. Test drugs: Comparative compound UA (urolithiasis A, 50 mg / kg) and example compounds CP-01 (Ua-a01 hydrochloride, 85.66 mg / kg), CP-10 (Ua-d01 hydrochloride, 85.66 mg / kg), and CP-18 (Ua-f01 hydrochloride, 121 mg / kg).

[0234] 2. Experimental animals: 20 male ICR mice, SPF grade.

[0235] 3. Drug administration and sample collection: Animals were fasted overnight (12h) before drug administration, with free access to water. They were weighed before drug administration, and the dosage was calculated based on their body weight. The drug was administered orally via gavage, and the animals were fed 4 hours after drug administration.

[0236] Blank blood was collected before drug administration, and blood was collected at predetermined time points after drug administration: 0.25h, 0.5h, 1h, 2h, 4h, 6h, 8h, and 24h. Blood was collected via the jugular vein or other suitable method, with 0.03mL of blood collected per sample. Heparin sodium was used for anticoagulation, and the samples were placed on ice after collection.

[0237] 4. Sample processing: After blood sample collection, place it on ice and centrifuge to separate plasma within 1 hour (centrifugation conditions: 6800g, 6min, 2-8℃). Plasma samples should be stored at -80℃ before analysis.

[0238] 5. Pharmacokinetic Analysis: Pharmacokinetic parameters, namely AUC, were calculated using Phoenix WinNonlin based on plasma drug concentration data at different time points. (0-∞) T 1 / 2 C max T max and MRT (0-∞) .

[0239] The specific test results are shown in Table 3. As can be seen from Table 3, all three compounds in this application exhibited superior pharmacokinetic properties compared to UA. Specifically, all three compounds in this application were absorbed and reached peak concentration relatively quickly, and their plasma concentrations and area under the curve (AUC) were significantly higher than those of UA, demonstrating better in vivo exposure characteristics and overcoming the problem of low bioavailability of UA.

[0240] Table 3 Pharmacokinetic parameters

[0241] Test Example 4: The therapeutic effect of compound CP-01 on Friedreich's ataxia

[0242] 1. Test drugs: Comparative compound UA (urolithin A) and example compound CP-01 (Ua-a01 hydrochloride).

[0243] 2. Experimental animals: SPF-grade YG8R mice (Fxntm1Mkn Tg(FXN)YG8Pook / J(YG8R) transgenic mice) and WT mice (C57BL / 6VAF / Elite mice).

[0244] 3. Animal grouping and administration: Mice were randomly divided into 7 groups, with 9–12 females and 9–13 males in each group. Mice were administered the drug via gavage at 6 months of age. The specific groupings and administration methods were as follows: normal control group (WT mice, saline), normal administration group (WT mice, CP-01, 36 mg / kg), model group (YG8R mice, saline), low-dose group (YG8R mice, CP-01, 18 mg / kg), medium-dose group (YG8R mice, CP-01, 36 mg / kg), high-dose group (YG8R mice, CP-01, 72 mg / kg), and positive control group (YG8R mice, UA, 172 mg / kg).

[0245] 4. Evaluation Methods: Mice in each group were administered the appropriate dose of the drug or saline via gavage for 22 weeks. Weight was measured every two weeks during this period. At 7, 8, 9, 10, and 11 months of age, mice underwent behavioral tests including grip strength, rotarod performance, and open field tests. The neuroprotective effect of compound CP-01 on YG8R mice was evaluated using these behavioral indicators. The specific details of each behavioral experiment are as follows:

[0246] Grasping force test: The gripping force test is mainly used to assess the limb muscle strength of animals. A mouse is gently placed in the center of a gripping force meter, and its tail is gently pulled to encourage the mouse to grasp the gripping plate with its limbs. Then, the mouse's tail is pulled backward at a constant speed until the mouse's forelimbs release the gripping plate. The maximum value displayed on the gripping force meter before the mouse releases the gripping plate is recorded. Each mouse is tested three times, with a 30-minute interval between each test. The maximum value from the three tests is taken as the evaluation value.

[0247] Roulette test: The roulette test is mainly used to assess the endurance, balance, and coordination of mice. During the training phase, mice were placed on a roulette fatigue tester with the rotation speed set to a maximum of 30 rpm. All mice were trained twice a day for 5 minutes each time, for a total of 3 days. The formal experiment was repeated 3 times, with at least 1 hour intervals between each test. The average time spent on the roulette for each mouse was recorded and calculated.

[0248] Open field test: The open field test is mainly used to observe the autonomous behavior of animals in novel environments and to explore their stress levels. In a quiet environment, mice are placed in the central area of ​​the test chamber, and their exploration routes and exploration time in different areas are recorded over 10 minutes. Before and after training each mouse, the open field chamber is wiped with alcohol (75% by volume) to remove the odor of the previous mouse and prevent it from affecting the results of subsequent experiments.

[0249] 5. The experimental results are as follows:

[0250] Figure 1 shows the effect of CP-01 on the body weight of FA model mice. A) Male mice, B) Female mice. One-way ANOVA or grouped t-test was used for multiple comparisons. All results are expressed as mean ± SEM. ** p<0.01, # p<0.05, ns = no statistical significance.

[0251] Figure 2 shows the effect of CP-01 on the gripping force of FA model mice. A) Male mice, B) Female mice; One-way ANOVA or grouped t-test for multiple comparisons were used. All results are expressed as mean ± SEM. *** p<0.001, # p<0.05, ### p<0.001.

[0252] Figure 3 shows the effect of CP-01 on the rotarod test of FA model mice. A) Fall time of male mice, B) Fall time of female mice, C) Fall distance of male mice, D) Fall distance of female mice. One-way ANOVA or grouped t-test for multiple comparisons were used. All results are expressed as mean ± SEM. * p<0.05, ** p<0.01, # p<0.05.

[0253] Figure 4 shows the effect of CP-01 on the open field test results of FA model mice. A) Movement distance of male mice, B) Movement distance of female mice, C) Average speed of male mice, D) Average speed of female mice, E) Number of times male mice entered the central area, F) Number of times female mice entered the central area, G) Movement time of male mice in the central area, H) Movement time of female mice in the central area. One-way ANOVA or multiple comparisons using paired t-tests were used. All results are expressed as mean ± SEM. ## p<0.01, ns = no statistical significance.

[0254] The test results show that, as shown in Figure 1A), compared with the normal group, the male mice in the model group had a decreased body weight; as shown in Figure 2, compared with the normal group, the mice in the model group had impaired limb strength; and as shown in Figure 3, compared with the normal group, the mice in the model group had decreased motor coordination and balance. After 22 weeks of treatment with CP-01 by gavage, as shown in Figure 2A), the YG8R male mice showed a trend of increasing body weight, and the impaired limb strength was significantly improved; as shown in Figures 3A) to C), the motor coordination and balance of the YG8R male mice significantly increased. In contrast, the control compound UA did not show any therapeutic effect in any of the experimental groups.

[0255] In conclusion, CP-01 can effectively improve the motor function of male YG8R mice, may have a neuroprotective effect on mice, and has the potential to treat Friedreich ataxia.

[0256] Test Example 5: The therapeutic effect of CP-01 on ALS

[0257] 1. Test drugs: Comparative compound UA (urolithin A) and example compound CP-01 (Ua-a01 hydrochloride).

[0258] 2. Experimental animals: 12 SPF-grade C57BL / 6J mice, SOD1 G93A Forty-eight mice, 11 weeks old.

[0259] 3. Animal grouping and administration: Mice were randomly divided into 5 groups of 12 mice each. After one week of acclimatization feeding, they were randomly assigned to groups and administered the drugs via gavage. The specific grouping and administration were as follows: normal control group (WT mice, saline), model group (SOD1...). G93A Mice, saline, CP-01 low-dose administration group (SOD1) G93A Mice, 18 mg / kg), CP-01 medium-dose administration group (SOD1) G93A Mice, 36 mg / kg, UA control group (SOD1) G93A Mice, 36 mg / kg).

[0260] 4. Assessment Methods: Administer the drug once daily for 6 consecutive weeks. At the end of the drug administration period, conduct behavioral tests including pole climbing, hanging, gripping, and spinning. Details of each behavioral test are as follows:

[0261] Pole Climbing Test: This test is primarily used to assess the motor and coordination abilities of an animal's limbs. A homemade wooden pole (50cm long, 1cm in diameter) is wrapped in gauze to increase friction. The pole is placed vertically against a horizontal table, and a mouse is placed headfirst at the top. The mouse crawls down the pole independently without external force, and the time taken to climb from the top to the bottom platform is recorded. A cutoff value of 20 seconds is used for each test; any result exceeding 20 seconds is recorded. The test is repeated three times for each mouse, and the average crawling time for each mouse is recorded and calculated.

[0262] Suspension test: The grip strength of mice was evaluated using an inverted grid suspension test. Each mouse was placed in the center of a square wire grid with sides of 21 cm (line width: 0.1 cm, spacing: 0.5 cm). The grid was tapped to induce the mouse to grip, and then the grid was slowly inverted. The suspension time of the mouse on the grid was recorded. After 6 weeks of treatment, all mice were trained twice a day for 3 consecutive days, with 90 seconds as the cutoff value. The experiment was repeated 3 times for each mouse, and the average suspension time was recorded and calculated.

[0263] Grip test: See Test Example 4 for experimental procedures.

[0264] Rotating rod experiment: See Test Example 4 for experimental procedures.

[0265] 5. The experimental results are as follows:

[0266] Figure 5 shows the effect of CP-01 on behavioral tests of ALS model mice: A) suspension test, B) rotarod test, C) grip test, and D) pole climbing test. All results are expressed as mean ± SEM. * p<0.05, ** p<0.01, *** p<0.005. As shown in Figure 5, compared with the control group, the model group mice had impaired limb coordination, and their grip strength and muscle strength were significantly reduced; after gavage treatment with CP-01, all the mice's abilities were significantly restored, and the effect was better than that of the control compound UA.

[0267] In summary, CP-01 improves SOD1 G93A The mice's motor function suggests that CP-01 has the potential to treat ALS.

[0268] Test Example 6: The therapeutic effect of CP-01 on MJD

[0269] 1. Test drug: Compound CP-01 (Ua-a01 hydrochloride) from the examples.

[0270] 2. Experimental animals: 24 SPF-grade C57BL / 6J mice (12 males and 12 females) and 72 MJD mice (36 males and 36 females), 6 months old.

[0271] 3. Animal grouping and administration: Mice were randomly divided into 4 groups, with 12 males and 12 females in each group. After one week of acclimatization feeding, the mice were administered the drug by gavage. The specific grouping and administration were as follows: normal control group (WT mice, saline), model group (MJD mice, saline), low-dose CP-01 administration group (MJD mice, 18 mg / kg), and medium-dose CP-01 administration group (MJD mice, 36 mg / kg).

[0272] 4. Assessment Methods: Administer the drug continuously for 7 months, once daily. At the end of the drug administration period, behavioral tests were conducted, including pole climbing, hanging, gripping, and spinning. The specific details of each behavioral test are as follows:

[0273] Pole climbing experiment: See Test Example 5 for experimental procedures.

[0274] Suspension test: See Test Example 5 for experimental procedures.

[0275] Grip test: See Test Example 4 for experimental procedures.

[0276] Rotating rod experiment: See Test Example 4 for experimental procedures.

[0277] 5. The experimental results are as follows:

[0278] Figure 6 shows the effects of CP-01 on behavioral tests in MJD model rats: A) Male pole climbing test, B) Female pole climbing test, C) Male grip strength test, D) Female grip strength test, E) Male suspension test, F) Female suspension test, G) Male rotisserie test, H) Female rotisserie test; all results are expressed as mean ± SEM. * p<0.05, ** p<0.01, *** p<0.005. As shown in Figure 6, compared with the control group, the model group mice had impaired limb coordination, and their grip strength and muscle strength were significantly reduced; after treatment with CP-01 by gavage, all of the mice's abilities were significantly restored.

[0279] In conclusion, CP-01 improved the motor function of MJD mice, indicating that CP-01 has the potential to treat MJD.

[0280] Test Example 7: The therapeutic effect of CP-10 on DMD

[0281] 1. Test drugs: Comparative compound UA (urolithin A) and example compound CP-10 (Ua-d01 hydrochloride).

[0282] 2. Experimental animals: 9 SPF-grade C57BL / 10J mice and 27 male Dmdem3Cd4 / Gpt mice, 26 weeks old.

[0283] 3. Animal grouping and administration: Mice were randomly divided into 4 groups of 9 mice each. After one week of acclimatization feeding, they were randomly assigned to groups and administered the drugs via gavage. The specific grouping and administration were as follows: normal control group (WT mice, saline), model group (Dmdem3Cd4 / Gpt mice, saline), CP-10 administration group (Dmdem3Cd4 / Gpt mice, 72 mg / kg), and UA control group (Dmdem3Cd4 / Gpt mice, 36 mg / kg).

[0284] 4. Assessment Methods: Administer the drug once daily for 8 consecutive weeks. At the end of the drug administration period, behavioral tests including pole climbing, endurance, and grip strength tests will be conducted. Details of each behavioral test are as follows:

[0285] Pole climbing experiment: See Test Example 5 for experimental procedures.

[0286] Endurance Test: The exercise endurance of mice was assessed by measuring the number of electric shocks received on an uphill treadmill. Exercise program settings: initial speed 9 cm / s, incline angle 5°, acceleration 3 cm / s / 12 min. Mice that could not keep up with the treadmill speed received an electric shock. A mouse was considered exhausted if it received 5 shocks / min for two consecutive minutes. The training phase lasted 8 weeks, 3 days a week. After the training phase, the formal experiment began, and the number of electric shocks was recorded.

[0287] Grip test: See Test Example 4 for experimental procedures.

[0288] 5. The experimental results are as follows:

[0289] Figure 7 shows the effect of CP-10 on behavioral tests of DMD model mice: A) pole climbing test, B) endurance test, and C) gripping test; all results are expressed as mean ± SEM. ** p<0.01, *** p<0.001. As shown in Figure 7, compared with the normal group, the model group mice had impaired limb balance, significantly decreased exercise endurance, and significantly decreased limb grip strength. After treatment with CP-10 by gavage, the mice's limb balance, endurance, and grip strength all significantly recovered to near the normal group. The UA gavage group also showed a trend of improvement, but it was not significant.

[0290] In conclusion, CP-10 can effectively improve the motor function of DMD model mice and has the potential to treat DMD.

[0291] Test Example 8: The therapeutic effect of CP-10 on ALS

[0292] 1. Test drugs: Comparative compound UA (urolithin A) and example compound CP-10 (Ua-d01 hydrochloride).

[0293] 2. Experimental animals: 9 SPF-grade C57BL / 10J mice, SOD1 G93A Twenty-seven mice, 11 weeks old.

[0294] 3. Animal grouping and administration: Mice were randomly divided into 4 groups of 9 mice each. After one week of acclimatization feeding, they were randomly assigned to groups and administered the drug orally. Specific grouping and administration details were as follows: normal control group (WT mice, saline), model group (SOD1...). G93A Mice, saline, CP-10 administration group (SOD1) G93A Mice, 72 mg / kg), UA control group (SOD1) G93A Mice, 36 mg / kg).

[0295] 4. Assessment Methods: Administer the drug once daily for 6 consecutive weeks. At the end of the drug administration period, behavioral tests such as pole climbing, hanging, and gripping were conducted. The specific details of each behavioral test are as follows:

[0296] Pole climbing experiment: See Test Example 5 for experimental procedures.

[0297] Suspension test: See Test Example 5 for experimental procedures.

[0298] Grip test: See Test Example 4 for experimental procedures.

[0299] 5. The experimental results are as follows:

[0300] Figure 8 shows the effect of CP-10 on behavioral tests of ALS model mice: A) pole climbing test, B) suspension test, and C) gripping test; all results are expressed as mean ± SEM. * p<0.05, ** p<0.01, *** p<0.005. As shown in Figure 8, compared with the control group, the model group mice had impaired limb coordination, and their grip strength and muscle strength were significantly reduced; after treatment with CP-10, all of the mice's abilities were significantly restored, and the effect was better than that of the control compound UA.

[0301] In summary, CP-10 improves SOD1 G93A The mice's motor function suggests that CP-10 has the potential to treat ALS.

[0302] Test Example 9: The therapeutic effect of CP-18 on ALS

[0303] 1. Test drugs: Comparative compound UA (urolithin A) and example compound CP-18 (Ua-f01 hydrochloride).

[0304] 2. Experimental animals: 20 SPF-grade WT mice (litter-bred mice without genetic diseases), SOD1 G93A Fifty mice, 13 weeks old.

[0305] 3. Animal grouping and administration: Mice were randomly divided into 7 groups of 10 mice each. Mice were administered the drug via gavage at 3 months of age. The specific grouping and administration details were as follows: normal control group (WT mice, saline), normal administration group (WT mice, CP-18, 100 mg / kg), and model group (SOD1). G93A Mice, saline, low-dose group (SOD1) G93A Mice, CP-18, 25 mg / kg, medium dose group (SOD1) G93A Mice, CP-18, 50 mg / kg, high-dose group (SOD1) G93AMice, CP-18, 100 mg / kg, and the model group given the positive drug (SOD1) G93A Mice, UA, 10 mg / kg).

[0306] 4. Assessment Methods: Each group was administered the appropriate dose of medication or saline via gavage for 6 weeks, followed by behavioral testing and pathological examination. Specific methods are as follows:

[0307] Rotating rod experiment: See Test Example 4 for experimental procedures.

[0308] Pole climbing experiment: See Test Example 5 for experimental procedures.

[0309] Grip test: See Test Example 4 for experimental procedures.

[0310] Suspension test: See Test Example 5 for experimental procedures.

[0311] Gait detection: The natural gait of mice was acquired through a gait tracking and analysis system, and gait detection and analysis were performed. The entire gait test was conducted in a quiet, dark environment. During the training phase, mice were repeatedly allowed to walk across a glass walkway (50cm long) to adapt to the environment. During the formal test, the mice were placed at the entrance of the walkway and made to walk forward. Walking data was collected using a high-speed gigabit Ethernet camera and filtered and analyzed by the system software WalkAnalysator. If the mouse paused or turned back during walking, the measurement had to be repeated.

[0312] Serum creatine kinase (CK) and lactate dehydrogenase (LDH) detection: Blood was collected via the inferior vena cava before euthanasia of mice, centrifuged (4°C, 1500g, 10min), and then CK and LDH were measured using an automated clinical analyzer (HITACHI 7080).

[0313] HE staining: Sections were routinely dewaxed to water, hematoxylin stained the nuclei for 5 min, tap water was used for blueing for 10 min, eosin stained for 3 min, and finally dehydrated with ethanol (95% v / v) for 10 s, followed by dehydration three times with anhydrous ethanol for 10 s each time; xylene was used for clearing three times for 5 min each time. The sections were then mounted with neutral resin.

[0314] Sirius red staining: Sections are routinely dewaxed to water, then iron-hematoxylin staining solution is added for 5–10 min, followed by rinsing with tap water for 10 min. After staining with Sirius red solution for 1 hour, the sections are rinsed slightly with running water to remove the stain. Then, sections are stained with 75% ethanol for 1 min, 95% ethanol for 1 min, anhydrous ethanol for 1 min, and xylene three times, 1–2 min each time, and mounted with neutral resin.

[0315] Masson staining: Sections were routinely dewaxed to water, stained with hematoxylin for 5–10 min, then differentiated with acidic ethanol differentiation solution, soaked in distilled water for 1 min to regain blue color, stained with Ponceau S and fuchsin for 10 min, washed with weak acid working solution for 1 min, washed with phosphomolybdic acid solution for 2 min, washed with weak acid working solution for 1 min, then stained with aniline blue staining solution for 2 min, washed with prepared weak acid working solution for 1 min. Finally, dehydrated with 95% ethanol for 10 s, dehydrated with anhydrous ethanol 3 times for 10 s each time; cleared with xylene 3 times for 5 min each time. Mounted with neutral resin.

[0316] Immunohistochemistry: Sections were routinely dewaxed to water, then immersed in antigen retrieval solution (1×), heated in a microwave oven on medium heat for 10 min, washed three times with PBS (pH=7.4) for 3 min each time; incubated with hydrogen peroxide blocking solution for 15 min; washed three times with PBS on a shaker for 10 min each time; incubated with protease blocking solution BSA (containing 0.3% Triton-100) for 1 h; incubated directly with the corresponding diluted primary antibody working solution at room temperature for 1 h; then transferred to a 4°C refrigerator overnight; removed the overnight sections and equilibrated at room temperature for 1 h, then washed four times with PBS on a shaker for 10 min each time; incubated with an appropriate amount of secondary antibody working solution for 1 h; washed three times with PBS on a shaker for 5 min each time; stained the treated sections in DAB working solution for 1–10 min; washed twice with PBS; dehydrated sequentially with 75%, 85%, 95%, and 100% ethanol for 3 min each time; cleared twice with xylene for 5 min each time; mounted with neutral resin.

[0317] Mitochondrial copy number detection: Total DNA was extracted from mouse gastrocnemius muscle tissue using a DNA extraction kit. The mitochondrial primer sequences were as follows:

[0318] F: 5'-gccagcctgacccatagccataat-3' (SEQ ID NO. 1);

[0319] R: 5'-gccggctgcgtattctacgtta-3' (SEQ ID NO. 2).

[0320] The sequence of the nuclear internal reference primer is as follows:

[0321] F: 5'-ttgagactgtgattggcaatgcct-3' (SEQ ID NO.3);

[0322] R: 5'-ccagaaatgctgggcgtact-3' (SEQ ID NO. 4).

[0323] Dilute the DNA with sterile water according to the specified concentration and prepare the qPCR reaction system as per the instructions. Then perform the qPCR test. The obtained data are calculated as follows:

[0324] Relative mitochondrial DNA content = 2 × 2 - ΔCT (ΔCT = mitochondrial CT - nuclear CT).

[0325] ATP and MDA level detection in tissues: ATP levels in the gastrocnemius muscle were detected using an ATP assay kit. Tissue samples were homogenized with ATP lysis buffer at 4°C, followed by centrifugation to remove the precipitate. A portion of the supernatant was used to determine protein concentration using a BCA assay kit. A reaction mixture of 50 μL of supernatant and 100 μL of ATP assay reagent was added to a 96-well black plate and incubated at room temperature for 3 min. ATP levels were then measured using a multi-mode microplate reader. For MDA detection, lipid peroxidation levels in the hippocampus were detected using a lipid peroxidation MDA assay kit. After quantifying the protein concentration, 100 μL of the extracted sample was mixed with 200 μL of MDA assay reagent and boiled at 100°C for 15 min, followed by centrifugation to collect the supernatant. 200 μL of the supernatant was added to a 96-well white plate, and absorbance was measured at 450 nm using a microplate reader.

[0326] 5. The experimental results are as follows:

[0327] The behavioral test results are shown in Figures 9, 10, and 11. Figure 9 shows the effect of CP-18 on the behavioral tests of ALS model mice: A) rotundus test, B) pole climbing test, C) suspension test, and D) gripping test; n = 8, and the data are expressed as mean ± SEM. * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001. Figure 10 shows the representative footprints in the gait test of CP-18 in ALS model mice. Figure 11 shows the effect of CP-18 on the limb coordination ability of ALS model mice. A) Average stride length of the left forelimb of mice, B) Average stride length of the right forelimb of mice, C) Average stride length of the left hindlimb of mice, D) Average stride length of the right hindlimb of mice; n=8, data are expressed as mean ± SEM. * p<0.05, ** p<0.01, ****p<0.0001. The results showed that, compared with the normal group, the model group mice had impaired limb balance, weakened limb strength, decreased grip strength (as shown in Figure 9), and decreased motor coordination and motor ability (as shown in Figures 10 and 11). After treatment with CP-18 by gavage, the mice's limb balance, limb strength, grip strength, motor ability, and coordination all recovered. The UA group by gavage also showed a trend of improvement, but there was no significant difference (as shown in Figures 9, 10, and 11).

[0328] Figure 12 shows the effect of CP-18 on muscle damage in ALS model mice. A) Serum creatine kinase (CK) level in mice, B) Serum lactate dehydrogenase (LDH) level in mice; n=8, data are expressed as mean ± SEM. * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001. Biochemical results showed that, compared with the normal group, the levels of creatine kinase and lactate dehydrogenase in the serum of the model group, which are related to muscle damage, were significantly increased; after administration of CP-18, the corresponding indicators decreased (as shown in Figure 12), indicating that CP-18 can improve muscle damage in model mice.

[0329] Figure 13 shows the effects of CP-18 on the inflammatory infiltration and fibrosis levels of the gastrocnemius muscle in ALS model mice. A) HE staining, B) Sirius red staining, C) Masson staining; the arrangement of the staining results in C) is consistent with the arrangement of the staining results in A) and B). HE staining, Masson staining, and Sirius red staining results show that, compared with the normal group, the inflammatory infiltration and fibrosis levels of the gastrocnemius muscle in the model group are increased; after oral administration of CP-18, the muscle inflammation and fibrosis levels of the model mice decreased (as shown in Figure 13).

[0330] The results of immunohistochemical (NeuN, CHAT, GFAP, IBA1) tests are shown in Figures 14, 15, 16, and 17. Figure 14 shows the effect of CP-18 on the number of spinal cord neurons in ALS model mice (immunohistochemical diagram), A) immunohistochemical diagram of anterior horn neurons in mouse spinal cord, B) immunohistochemical diagram of cholinergic neurons in anterior horn of mouse spinal cord; Figure 15 shows the effect of CP-18 on the number of spinal cord neurons in ALS model mice (statistical diagram), A) statistical diagram of the number of anterior horn neurons in mouse spinal cord, B) statistical diagram of the number of cholinergic neurons in anterior horn of mouse spinal cord; n=3, data are expressed as mean ± SEM; * p<0.05, **p<0.01, ***p<0.001. Figure 16 shows the effect of CP-18 on the level of spinal cord nerve inflammation in ALS model mice (immunohistochemical diagram), A) immunohistochemical diagram of Iba1, a marker of microglia activation in the anterior horn of the mouse spinal cord, B) immunohistochemical diagram of GFAP, a marker of astrocyte activation in the anterior horn of the mouse spinal cord; Figure 17 shows the effect of CP-18 on the level of spinal cord nerve inflammation in ALS model mice (statistical diagram), A) statistical diagram of the number of microglia in the anterior horn of the mouse spinal cord, B) statistical diagram of the number of astrocytes in the anterior horn of the mouse spinal cord; n=3, data are expressed as mean ± SEM; * p<0.05, ** p<0.01, *** p<0.001. The results showed that, compared with the control group, the number of motor neurons in the model group mice was significantly reduced, and the number of activated astrocytes and microglia was significantly increased; after CP-18 treatment, the number of neurons in the model group mice increased and the number of glial cells decreased (as shown in Figures 14, 15, 16 and 17), indicating that CP-18 can improve neuronal loss in mice and has a certain therapeutic effect on neuroinflammation.

[0331] Figure 18 shows the effects of CP-18 on the number and function of mitochondria in the gastrocnemius muscle of ALS model mice. A) Mitochondrial copy number in mouse gastrocnemius muscle, B) ATP level in mouse gastrocnemius muscle, C) MDA content in mouse gastrocnemius muscle; n=5, data are expressed as mean ± SEM. * p<0.05, ** p<0.01, **** p<0.0001. As shown in Figure 18, compared with the normal group, the number of mitochondria in the model group mice was reduced and the mitochondrial function was significantly decreased; after treatment with CP-18, the number of mitochondria in the model mice was significantly increased and the mitochondrial function was significantly enhanced. The UA gavage group also showed a trend of improvement, but it was not significant.

[0332] In conclusion, CP-18 can effectively improve SOD1 G93A The study showed that CP-18 significantly improved the motor function of the model mice, as well as its therapeutic effects on muscle inflammation and fibrosis, spinal cord inflammation, and neuronal loss. It also improved mitochondrial function in the mice. This suggests that CP-18 has potential therapeutic effects on ALS, and that this efficacy may be related to the improvement of mitochondrial function.

[0333] Test Case 10: Anti-aging effects of CP-18 and its therapeutic effects on sarcopenia

[0334] 1. Test drugs: Comparative compound UA (urolithin A) and example compound CP-18 (Ua-f01 hydrochloride).

[0335] 2. Experimental animals: 20 SPF-grade C57BL / 6J mice, 2 months old; 50 SPF-grade C57 mice, 18.5 months old.

[0336] 3. Animal grouping and administration: Mice were randomly divided into 7 groups of 10 mice each. Administered the medication via gavage. The specific groupings and administration methods were as follows: Young control group (young C57 mice, saline), Young administration group (young C57 mice, CP-18, 100 mg / kg), Natural aging group (aged C57 mice, saline), Low-dose group (aged C57 mice, CP-18, 25 mg / kg), Medium-dose group (aged C57 mice, CP-18, 50 mg / kg), High-dose group (aged C57 mice, CP-18, 100 mg / kg), and UA control group (aged C57 mice, UA, 10 mg / kg).

[0337] 4. Assessment method: Each group was given the corresponding dose of drug or saline by gavage for 10 weeks, followed by behavioral tests and pathological examinations.

[0338] Specifically as follows:

[0339] Rotating rod experiment: See Test Example 4 for experimental procedures.

[0340] Pole climbing experiment: See Test Example 5 for experimental procedures.

[0341] Grip test: See Test Example 4 for experimental procedures.

[0342] Suspension test: See Test Example 5 for experimental procedures.

[0343] Gait detection: See Test Example 9 for experimental procedures.

[0344] HE staining: See Test Example 9 for experimental procedures.

[0345] Masson staining: See Test Example 9 for experimental procedures.

[0346] Immunohistochemistry: See Test Example 9 for experimental procedures.

[0347] Detection of ATP and MDA levels in tissues: See Test Example 9 for experimental procedures.

[0348] Mitochondrial copy number and telomere length determination: Total DNA was extracted from mouse gastrocnemius muscle tissue using a DNA extraction kit. The primer sequences for mitochondria are shown in SEQ ID NO.1 and SEQ ID NO.2.

[0349] The primer sequences for the nuclear internal reference are shown in SEQ ID NO.3 and SEQ ID NO.4.

[0350] Telomere primer sequences:

[0351] F: 5'-ggtttttgagggtgagggtgagggtgagggtgagggt-3' (SEQ ID NO. 5);

[0352] R: 5'-tcccgactatccctatccctatccctatccctatcccta-3' (SEQ ID NO. 6);

[0353] The primer sequences for the single-copy gene are as follows:

[0354] F: 5'-cagcaagtgggaaggtgtaatcc-3' (SEQ ID NO.7);

[0355] R: 5'-cccattctatcatcaacgggtacaa-3' (SEQ ID NO. 8).

[0356] Dilute the DNA with sterile water according to the DNA concentration, prepare the qPCR reaction system according to the instructions, and then perform the instrument detection.

[0357] 5. The experimental results are as follows:

[0358] The behavioral test results are shown in Figures 19, 20, and 21. Figure 19 shows the effects of CP-18 on the motor function and muscle strength of naturally aging mice. A) Rotary bar test, B) Pole climbing test, C) Suspension test, D) Grasp test; n=8, data are expressed as mean ± SEM. * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001. Figure 20 shows representative footprints in the gait test of naturally aged mice by CP-18; Figure 21 shows the effect of CP-18 on the limb coordination ability of naturally aged mice, A) average stride length of mouse limbs, B) statistics of mouse movement speed, C) statistics of mouse gait frequency, D) statistics of mouse footprint cycle time; n=8, data are expressed as mean ± SEM; * p<0.05, ** p<0.01, **** p<0.0001. The results showed that compared with the young group, the aged mice had impaired limb balance, weakened limb strength, decreased grip strength (as shown in Figure 19), and decreased motor coordination and motor ability (as shown in Figures 20 and 21). After treatment with CP-18 by gavage, the limb balance, limb strength, grip strength, motor ability, and coordination of the aged mice were restored. The UA group by gavage also showed a trend of improvement, but it was not significant (as shown in Figures 19, 20, and 21).

[0359] Figure 22 shows the effects of CP-18 on the levels of inflammatory infiltration and fibrosis in the gastrocnemius muscle of naturally aging mice. A) HE staining, B) Masson staining, C) Immunohistochemical staining of Fibronectin protein; the arrangement of the staining results in B) is consistent with the arrangement of the staining results in A) and C). The results of HE staining, Masson staining, and immunohistochemistry of fibrosis protein showed that, compared with the young group, the aging group of mice had higher levels of inflammatory infiltration and fibrosis in the gastrocnemius muscle, and oral administration of CP-18 could improve the levels of muscle inflammation and fibrosis in aging mice (as shown in Figure 22).

[0360] Figure 23 shows the effect of CP-18 on mitochondrial function of gastrocnemius muscle in naturally aged mice. A) ATP level, B) MDA content; n=4, data are expressed as mean ± SEM. * p<0.05, ** p<0.01. As shown in Figure 23, compared with the young group, the ATP level in the gastrocnemius muscle of the aging group mice was significantly decreased and the lipid oxidation level was increased; the mitochondrial function of the aging group mice was enhanced after administration of CP-18; although the UA group also showed a trend of improvement, it was not significant.

[0361] Figure 24 shows the effects of CP-18 on mitochondrial copy number and telomere length in the gastrocnemius muscle of naturally aged mice. A) Mitochondrial copy number, B) Telomere length; n=5, data are expressed as mean ± SEM. * p<0.05, ** p<0.01. The results of gastrocnemius muscle mitochondrial number and telomere length detection showed that, compared with the young group, the number of gastrocnemius muscle mitochondrial number and telomere length of aged mice were reduced, while the number of mitochondrial number and telomere length increased after administration of CP-18 (as shown in Figure 24).

[0362] Figure 25 shows the effects of CP-18 on the appearance of naturally aging mice. The results showed that compared with the younger group, the aging group mice had hair loss on their backs and tails, messy fur, and more white hair. After oral administration of CP-18, the hair loss on the backs and tails of the mice was improved, the fur was shinier and softer, and the white hair was reduced.

[0363] Figure 26 shows the effect of CP-18 on the aging of the gastrocnemius muscle in naturally aging mice (representative image of the right hind limb of the mouse), and Figure 27 shows the effect of CP-18 on the aging of the gastrocnemius muscle in naturally aging mice (statistical graph of gastrocnemius muscle wet weight ratio). The results show that compared with the young group, the wet weight ratio of the gastrocnemius muscle in the aging group decreased significantly, while the wet weight ratio of the gastrocnemius muscle in mice treated with CP-18 increased, indicating that CP-18 has an anti-atrophy effect on muscles.

[0364] Figure 28 shows the effects of CP-18 on cognitive impairment in naturally aging mice. A) Cognitive index statistics in the new object recognition test; B) Freezing index statistics in the scene-conditioned fear memory test; C) Freezing index statistics in the scene-conditioned fear memory test. The results showed that compared with the younger group, the cognitive index of aging mice in the new object recognition test was significantly decreased. The number of freezing points in the scene-conditioned fear memory test did not change significantly in aging mice, but the number of freezing points in the conditioned fear memory test was reduced. After CP-18 treatment by gavage, the cognitive index of aging mice increased, suggesting an increase in the number of freezing points in the conditioned fear memory test. This indicates that CP-18 can improve cognitive impairment in aging mice.

[0365] Figures 29, 30, 31, and 32 show the results of CP-18 on the expression of serum proteases in naturally aging mice. Figure 29 is a clustering diagram of differentially expressed proteins upregulated after CP-18 administration; Figure 30 is a heatmap of Cluster 1; Figure 31 is a clustering diagram of differentially expressed proteins downregulated after CP-18 administration; and Figure 32 is a heatmap of Cluster 2. The results show that CP-18 treatment improved the expression of serum proteases in aged mice, significantly reversing the expression of multiple aging-related proteins. The expression levels of differentially expressed proteins in aged mice were closer to those in younger mice, further suggesting that CP-18 has an anti-aging effect.

[0366] In conclusion, CP-18 can enhance and improve the motor function of aging mice, significantly improve muscle inflammation and fibrosis in aging mice, and also increase the number, function, and telomere length of mitochondria in the gastrocnemius muscle of aging mice, indicating that CP-18 has the potential to significantly improve aging and treat sarcopenia.

[0367] Test Example 11: The therapeutic effect of CP-18 on Pompe disease

[0368] 1. Test drug: Comparative compound UA and example compound CP-18 (Ua-f01 hydrochloride).

[0369] 2. Experimental animals: SPF-grade GAA knockout mice and mice from the same litter without genetic diseases; 12 months old.

[0370] 3. Animal grouping and administration: Mice were randomly divided into 4 groups, with 8–9 mice in each group. The treatment groups were administered the drug via gavage at 12 months of age at a concentration of 5 mg / mL in physiological saline, with a dosage of 0.1 mL / 10 g. The specific groupings and administration methods were as follows: normal control group (WT mice, physiological saline), normal treatment group (WT mice, CP-18), model group (GAA knockout mice, physiological saline), and model treatment group (GAA knockout mice, CP-18).

[0371] 4. Assessment Methods: Each group was administered the appropriate dose of medication or saline via gavage for 4 weeks, followed by behavioral testing. Details are as follows:

[0372] Pole climbing experiment: See Test Example 5 for experimental procedures.

[0373] Rotating rod experiment: See Test Example 4 for experimental procedures.

[0374] Open field experiment: See Test Example 4 for experimental procedures.

[0375] Tail suspension test: Each mouse was suspended upside down at a height of about 40cm above the ground for 5 minutes. During the test, the number of times the mouse raised its head and the time it remained still were carefully observed and recorded.

[0376] After the behavioral experiment, the mice were deeply anesthetized with 1% pentobarbital and the following tissue samples were taken: the gastrocnemius muscle and myocardium of the mice were dehydrated, fixed, and embedded in paraffin to prepare sections with a thickness of 5 μm. The morphology of the gastrocnemius muscle and myocardium was observed under a 20x microscope by immunofluorescence staining.

[0377] 5. The experimental results are as follows:

[0378] Figure 33 shows the effect of CP-18 on behavioral tests of PD (Pompe disease) model mice. A) Pole climbing test, B) Rotary bar test, C) Movement distance of mice in open field test, D) Time spent by mice in the central area of ​​open field test, E) Number of times mice raised their heads in tail suspension test; all results are expressed as mean ± SEM. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001. As shown in Figures 33A) to B), compared with the normal group, the model group mice showed a significant decrease in limb strength and motor coordination; as shown in Figures 33A) to C), after CP-18 administration, the GAA knockout mice showed a significant improvement in limb strength, a significant increase in motor coordination and balance, and a significant increase in movement distance.

[0379] Figure 34 shows the effect of CP-18 on the expression of autophagy and lysosome-related proteins (Lamp1 and LC3) in PD model mice (gastrocnemius muscle stained section), and Figure 35 shows the effect of CP-18 on the expression of autophagy and lysosome-related proteins (Lamp1 and LC3) in PD model mice (myocardial stained section). Immunofluorescence staining results of the gastrocnemius and myocardium of mice showed that, compared with WT mice, the autophagy-related protein markers (LC3) and lysosome markers (LAMP1) were significantly increased in the model mice, while LC3 and LAMP1 decreased after CP-18 treatment. Furthermore, compared with WT mice, the colocalization of LC3 and LAMP1 was reduced in the model mice. This indicates that CP-18 treatment improved the accumulation of autophagosomes and lysosomes in the model mice.

[0380] In conclusion, CP-18 can effectively improve the motor function of GAA knockout mice and improve their autophagosome and lysosomal levels, indicating that CP-18 has potential therapeutic efficacy for Pompe disease.

[0381] The above description is only a preferred embodiment of this application. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of this application, and these improvements and modifications should also be considered within the scope of protection of this application.

Claims

1. A urolithin A derivative or a pharmaceutically acceptable salt thereof, said urolithin A derivative having the structure shown in Formula 1: In Formula 1, X1 and X2 are independently selected from -O-, -S-, -NR3- or carbonyl, and at least one of X1 and X2 is carbonyl; R1 is selected from -H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, -C(=O)R4, -C(=O)OR5 or -C(=O)NR6R7. It consists of 3- to 10-membered nitrogen-containing heterocycles; n is an integer from 0 to 18; R2 is independently selected from deuterium, halogen, amino, nitro, carbonyl, ester, carboxyl, sulfonic acid, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C1-C6 amino, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocyclic alkyl; or R2 together with the attached nitrogen-containing heterocycle forms a substituted or unsubstituted spirocyclic ring or a substituted or unsubstituted bridged ring; R3, R4, R5, R6, and R7 are independently selected from -H, deuterium, substituted or unsubstituted C1-C6 chain alkyl, substituted or unsubstituted C3-C7 cycloalkyl, substituted or unsubstituted C3-C7 heterocyclic alkyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl; or R6 and R7 together with the attached nitrogen atom form a substituted or unsubstituted C3-C7 heterocyclic alkyl.

2. The urolithin A derivative or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, X1 and X2 are -O- and carbonyl groups, respectively.

3. The urolithin A derivative or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, The R1 is selected from -H, -CH3, -C(=O)CH3, -C(=O)OC(CH3)3, -C(=O)N(CH3)2 or 4. The urolithin A derivative or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, The R2 is selected from -CH3 or -N(CH3)2, or R2 together with the attached nitrogen-containing heterocycle forms an unsubstituted spiro ring or an unsubstituted bridged ring.

5. The urolithin A derivative or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, The Selected from 6. The urolithin A derivative or a pharmaceutically acceptable salt thereof according to any one of claims 2 to 5, characterized in that, The Selected from 7. The urolithin A derivative or a pharmaceutically acceptable salt thereof according to claim 6, characterized in that, The urolithin A derivative is selected from any one of the following compounds:

8. The urolithin A derivative or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, Pharmaceutically acceptable salts of the urolithin A derivatives include hydrochloride, sulfate, phosphate, hydrobromide, nitrate, salicylate, benzoate, C1-C6 fatty carboxylates, C1-C6 alkyl sulfonates, benzene sulfonates, p-toluene sulfonate, or camphor sulfonate.

9. A method for preparing the urolithin A derivative according to any one of claims 1 to 8, comprising the following steps: (I) When R1 is -H, and the When the urolithin A derivative is a nitrogen-containing heterocycle that does not contain a -NH- group, the preparation method includes the following steps: Compound a, compound b, a base reagent, and an organic solvent were mixed and subjected to a substitution reaction to obtain compound c. Compound c was subjected to a hydroxyl deprotection reaction to obtain compound d, wherein compound d is a compound with R1 being -H and The urolithin A derivative is a nitrogen-containing heterocycle that does not contain a -NH- group; The structural formulas of compounds a, b, and c are shown below in sequence: Z is a hydroxyl protecting group, X1, X2, R2 and n are defined as in Equation 1; (II) When R1 is -H, and the When the urolithin A derivative is a nitrogen-containing heterocycle containing a -NH- group, the preparation method includes the following steps: Compound a, compound b', a base reagent and an organic solvent were mixed and subjected to a substitution reaction to obtain compound c'; Compound c' is subjected to a hydroxyl deprotection reaction and an imine deprotection reaction to yield compound e, wherein compound e is R1-H and The urolithin A derivative is a nitrogen-containing heterocycle containing a -NH- group; The difference between compound b' and compound b is that compound b' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle; the difference between compound c' and compound c is that compound c' contains an imino protecting group at the -NH- group corresponding to the nitrogen-containing heterocycle. (III) When R1 is not -H, the preparation method of the urolithin A derivative includes the following steps: Compound d was prepared according to (I) or compound e was prepared according to (II); In the presence of an alkaline reagent and an organic solvent, compound d or compound e is subjected to a substitution reaction with compound f to obtain the urolithin A derivative in which R1 is not -H; The compound f is R4C(=O)X3, R5OC(=O)X3, R6R7NC(=O)X3 or R8X3, where X3 is a halogen atom and R8 is a substituted or unsubstituted C1-C6 chain alkyl, a substituted or unsubstituted C3-C7 cycloalkyl, a substituted or unsubstituted C3-C7 heterocycloalkyl, a substituted or unsubstituted aryl, or a substituted or unsubstituted heteroaryl.

10. The use of the urolithin A derivative or a pharmaceutically acceptable salt thereof as described in any one of claims 1 to 8 in the preparation of a medicament for the prevention and / or treatment of diseases related to mitochondrial function.

11. The application according to claim 10, characterized in that, Diseases associated with mitochondrial function include neurological disorders, aging, muscle diseases, or metabolic diseases.

12. The application according to claim 11, characterized in that, The neurological disorders mentioned include Friedreich's ataxia, amyotrophic lateral sclerosis (ALS), adrenoleukodystrophy, Alexander disease, Alpert disease, familial fatal insomnia, Huntington's disease, Kennedy's disease, Crabbe's disease, Lyme disease, Machado-Joseph disease, multiple sclerosis, multiple system atrophy, Niemann-Pick disease, Pick's disease, primary lateral sclerosis, progressive supranuclear palsy, Rafe-Syme disease, Sandhoff's disease, diffuse demyelinating sclerosis, spinocerebellar ataxia, subacute mixed degeneration of the spinal cord, toxic encephalopathy, disseminated cavernous encephalopathy, or ataxia-telangiectasia.

13. The application according to claim 11, characterized in that, The muscle diseases mentioned include sarcopenia or Duchenne muscular dystrophy.

14. The application according to claim 11, characterized in that, The metabolic diseases mentioned include obesity, hyperlipidemia, or Pompe disease.

15. A pharmaceutical composition comprising at least one of the urolithin A derivatives of any one of claims 1 to 8 and a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

16. The pharmaceutical composition according to claim 15, characterized in that, The pharmaceutical composition contains 0.1 to 99.9 wt% urolithiasis A derivatives and / or their pharmaceutically acceptable salts.

17. A method for preventing and / or treating diseases related to mitochondrial function, comprising administering to a person preventing and / or treating a disease related to mitochondrial function an effective amount of at least one of the urolithin A derivatives of any one of claims 1 to 8 and pharmaceutically acceptable salts thereof, or the pharmaceutical composition of claim 15 or 16.

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