Nucleoside derivatives for the treatment of cancer and use thereof

Abstract

The application discloses a nucleoside compound with a novel structure, a salt of the nucleoside compound and a preparation method of the nucleoside compound. The nucleoside compound and the salt thereof exhibit a strong inhibitory effect on tumor proliferation in vivo, and do not exhibit obvious side effects. Meanwhile, the compound of the application does not have obvious stomach irritation and kidney toxicity, and shows a good medical application prospect in the treatment of cancer.

Description

Technical Field

[0001] This invention relates to a class of compounds with anticancer activity, pharmaceutical compositions, and methods for synthesizing such compounds. Furthermore, this invention provides compositions and combinations of nucleoside compounds, and their roles in cancer treatment. Background Technology

[0002] Cancer is a disease that seriously endangers human life and health. Clinical manifestations mainly include fever, weight loss, insomnia, infection, abdominal pain, headache, and limb weakness. According to the WHO (World Health Organization), the number of new cancer cases worldwide reached 20 million in 2020, with approximately 10 million deaths. Cancer may still be the leading cause of death in this century, seriously affecting human physical and mental health and even posing the most serious threat to human survival.

[0003] Currently, cancer treatment mainly relies on chemotherapy. There are many types of anticancer drugs, with significant differences in their mechanisms of action, indications, and clinical applications. The main mechanisms of anticancer drugs include: (1) inhibiting cell proliferation by blocking the synthesis of ribonucleic acid (RNA), deoxyribonucleic acid (DNA), or proteins, thereby inducing apoptosis in cancer cells; (2) inhibiting cancer cell proliferation by altering hormone levels. Nucleoside analogs account for a large proportion of anticancer drugs. They mainly utilize the principle of electron isosterism, chemically modifying the nucleoside unit substances required for DNA replication, phosphorylating them within the cell, inhibiting the synthesis of nucleoside triphosphates (NTPs), interfering with DNA or RNA molecule replication, competitively inhibiting DNA polymerase, and preventing cancer cell proliferation, ultimately leading to apoptosis in cancer cells. Currently, the main types of nucleoside anticancer drugs on the market are as follows: (1) purine analogs such as thioguanine (6-TG) and azathioprine, which can inhibit the conversion process of hypoxanthine and can be used to treat leukemia, but there is currently little research on these drugs; (2) guanine drugs such as nucleoside analogs nelarabine, which is a prodrug of 9-β+D-argifuramic acid guanine, accumulates faster and more in T cells than in B cells, and has a stronger inhibitory effect on T cells. It can be used for chemotherapy ineffective or relapsed cases. (2) Acute T-cell lymphoblastic lymphoma has a significant therapeutic effect, but the drug can cause significant chest pain and even has central nervous system toxicity; (3) Adenosine analogs such as fludarabine and clofarabine are mainly used to treat B-cell lymphoblastic leukemia, but their therapeutic effect on various cancers is poor; (4) Uracil or uridine drugs, such as 5-fluorouracil, mainly interfere with DNA synthesis by inhibiting thymidine nucleotide synthase, thereby blocking the conversion of deoxyuridine nucleotides to deoxythymidine nucleotides. However, these drugs have poor selectivity and can cause serious gastrointestinal diseases; (5) Cytidine analogs such as cytarabine have been extensively studied and have multiple varieties on the market. They mainly inhibit the synthesis of cancer cell DNA and induce cancer cell apoptosis by converting to cytarabine triphosphate. However, these drugs are prone to causing side effects such as bone marrow suppression and thrombocytopenia.

[0004] Nucleoside analogs have diverse structures and exert their anticancer effects through different mechanisms. They remain a hot topic in anticancer drug research. However, there is still a need to develop more ideal nucleoside anticancer drugs. This requires not only in-depth research on improving efficacy, bioavailability, and reducing adverse reactions, but also breakthroughs in drug transport, pharmacokinetics, drug metabolism, and targeted therapy. Summary of the Invention

[0005] One object of the present invention is to provide a series of nucleoside compounds with novel structures and their salts, as well as methods for preparing such compounds.

[0006] Another objective of this invention is to provide compounds and compositions thereof that have a significant inhibitory effect on tumor proliferation in vivo and do not exhibit obvious toxic side effects.

[0007] The third objective of this invention is to provide compounds and compositions thereof that do not have significant gastric irritation and do not have significant nephrotoxicity, achieving unexpected results.

[0008] Unless otherwise specified herein, the technical terms used in this invention have basic meanings generally understood by those skilled in the art.

[0009] Furthermore, the compound provided by the present invention has the structure of formula (I):

[0010]

[0011] Or a pharmaceutically acceptable salt, solvate, isotope derivative, enantiomer, or diastereomer thereof, wherein:

[0012] W is selected from -O-, or does not exist;

[0013] m can be selected from 1, 2, 3, or 4;

[0014] R1 is selected from hydrogen, deuterium, fluorine, hydroxymethyl, hydroxyethyl, phenyl, C1-C6 linear or branched alkyl groups;

[0015] R2 is selected from hydrogen, deuterium, fluorine, cyano, hydroxymethyl, hydroxyethyl, phenyl, C1-C6 linear or branched alkyl, C2-C6 linear or branched alkenyl, and C2-C6 linear or branched alkynyl.

[0016] R3 is selected from hydrogen, hydroxyl group, fluorine, -OR5, and -O(C=O)R5;

[0017] R4 is selected from hydrogen, C1-C6 linear or branched alkyl, -(C=O)R5; wherein the alkyl group may be independently and optionally substituted by one or more substituents selected from the following: hydrogen, halogen, cyano, hydroxyl, carboxyl, trifluoromethyl, C1-C6 linear or branched alkoxy, C1-C6 linear or branched alkylamino.

[0018] R5 is selected from C1-C6 linear or branched alkyl, C3-C6 cycloalkyl, and C6-C15 aryl; wherein the alkyl, cycloalkyl, and aryl groups may be independently and optionally substituted by one or more substituents selected from the following: hydrogen, halogen, cyano, amino, hydroxyl, carboxyl, nitro, and trifluoromethyl.

[0019] When W is selected from -O-, L is selected from hydrogen, -(C=O)R5, or directly combined with R4 to form a group in the form of the following structural formula (I-1):

[0020]

[0021] When W does not exist, L is selected.

[0022] n is selected from 1 or 2;

[0023] Y is selected from -(C=O)-, or does not exist;

[0024] R a R b Each is independently selected from hydrogen, deuterium, C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, or R. a R b Connected to form a ring;

[0025] R6 is selected from hydrogen, C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, C3-C6 cycloalkyl, C6-C15 aryl, 3-membered to 6-membered ring monocyclic or bicyclic heterocyclic groups, -CH2O(C=O)R 11 -CH2O(C=O)OR 11 The alkyl, alkoxy, cycloalkyl, aryl, and heterocyclic groups therein may be independently and optionally substituted by one or more substituents selected from the following: hydrogen, halogen, cyano, trifluoromethyl, C1-C16 linear or branched alkyl, and C1-C16 linear or branched alkoxy.

[0026] R7 and R8 are each independently selected from hydrogen, C1-C6 linear or branched alkyl, C3-C6 cycloalkyl, C6-C15 aryl, (C1-C6 linear or branched alkyl)C6-C15 aryl, 3- to 6-membered monocyclic or bicyclic heterocyclic groups, and -CH2O(C=O)R. 11 -CH2O(C=O)OR 11 R7 and R8 are linked together to form a ring; wherein the alkyl, cycloalkyl, aryl, and heterocyclic groups may be independently and optionally substituted by one or more substituents selected from the following: hydrogen, halogen, cyano, trifluoromethyl, C1-C16 linear or branched alkyl, C1-C16 linear or branched alkoxy, C1-C6 linear or branched alkylamino.

[0027] R9 is selected from hydrogen, C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, C3-C6 cycloalkyl, C6-C15 aryl, 3-membered to 6-membered ring monocyclic or bicyclic heterocyclic groups, and -CH2O(C=O)R. 11 -CH2O(C=O)OR 11The alkyl, alkoxy, cycloalkyl, aryl, and heterocyclic groups therein may be independently and optionally substituted by one or more substituents selected from the following: hydrogen, halogen, cyano, trifluoromethyl, C1-C16 linear or branched alkyl, and C1-C16 linear or branched alkoxy.

[0028] R 10 The group is selected from C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, C1-C6 linear or branched alkylamino, C2-C6 linear or branched alkenyl, C2-C6 linear or branched alkynyl, C3-C6 cycloalkyl, C6-C15 aryl, 3- to 6-membered monocyclic or bicyclic heterocyclic groups, and 5- and / or 6-membered monocyclic or bicyclic heteroaryl groups; wherein the alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl, aryl, and heteroaryl groups may be independently and optionally substituted by one or more substituents selected from the following: hydrogen, halogen, cyano, amino, hydroxyl, carboxyl, nitro, trifluoromethyl, C1-C16 linear or branched alkyl, C1-C16 linear or branched alkoxy, C1-C6 linear or branched alkylamino, and C3-C6 cycloalkyl;

[0029] G is selected from

[0030] R 11 Selected from hydrogen, or -(C=O)R5;

[0031] W is selected from -O-, R 11 When L is selected from hydrogen or -(C=O)R5, L is selected from the form that directly combines with R4 to form a group with the following structural formula (I-1):

[0032]

[0033] W does not exist, R 11 When L is selected from hydrogen or -(C=O)R5, L is selected from...

[0034] R 12 Selected from hydrogen, halogen, hydroxyl, phenyl, and pyridyl;

[0035] R 13 Selected from hydrogen, C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, cyano, trifluoromethyl, phenyl, pyridyl;

[0036] R 14 It is selected from hydrogen, C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, cyano, and trifluoromethyl.

[0037] In a further aspect, compounds of formula (II) are provided.

[0038]

[0039] In equation (II), the definitions of each generation base are as defined in equation (I) above.

[0040] In a further aspect, compounds of formula (III) are provided.

[0041]

[0042] The substituents in equation (III) are defined as defined in equation (I) above.

[0043] In a further aspect, a compound is provided, selected from the following compounds:

[0044]

[0045]

[0046]

[0047] In a further aspect, the compound can be used in methods for preventing or treating cancers in humans or other mammals.

[0048] In a further aspect, the method wherein the cancer includes leukemia, pancreatic cancer, prostate cancer, lung cancer, liver cancer, stomach cancer, colorectal cancer, prostate cancer, bone marrow cancer, thyroid cancer, lymphoma, breast cancer, cervical cancer, ovarian cancer, and other related cancers.

[0049] In a further aspect, the compound is used in combination with other anticancer therapies or anticancer drugs.

[0050] In a further aspect, the use of the compound in the preparation of a medicament for the prevention or treatment of cancer in humans or other mammals.

[0051] In a further aspect, a pharmaceutical composition comprising the compound in combination with a pharmaceutical carrier, diluent, or excipient.

[0052] In a further aspect, a method of treating cancer comprises administering a therapeutically effective amount of any one of the compounds to a human or other mammal.

[0053] The terms “compound of formula (I)”, “compound of the present invention” or similar terms used above and below include compounds of formula (I) and any subgroup of compounds of formula (I), including all possible stereoisomers, pharmaceutically acceptable salts, solvates, isotope derivatives, and metal complexes.

[0054] The term "pharmaceutically acceptable salt" as used above and below refers to acid salts that can be obtained by treating the compounds of the present invention with suitable acids, including organic and inorganic acid salts. Organic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, etc.; organic acids include acetic acid, lactic acid, pyruvic acid, oxalic acid, succinic acid, tartaric acid, malic acid, methanesulfonic acid, p-toluenesulfonic acid, etc. Similarly, pharmaceutically acceptable metal salts or salts formed by combining with amines, such as sodium salts, potassium salts, magnesium salts, zinc salts, iron salts, calcium salts, ammonium salts, etc., can be obtained by treating with suitable organic or inorganic bases, and these are all included within the scope of the present invention.

[0055] The absolute configuration of the terms "enantiomer" and "diastereomer" used above and below can be determined using characterization methods known in the art, such as X-ray diffraction and NMR (Nuclear Magnetic Resonance). Preferably, the compositions of the present invention comprise stereoisomers of a single configuration of the above-described compound. "Single-configuration stereoisomer" means that the molecule substantially does not contain other enantiomers or diastereomers; specifically, the molecule contains at least 90% of the same isomer and at most 10% of other isomeric forms.

[0056] The stereoisomers of the compounds and their intermediates provided by this invention can be obtained by methods well known in the art, such as separation using preparative chromatography instruments, like a chromatographic column with a chiral stationary phase. Alternatively, they can be obtained by selectively crystallizing, separating, dissociating, and purifying the compounds and their intermediates using acids or bases with optical activity to form salts. For example, the acids used are L-tartaric acid, L-malic acid, S-mandelic acid, L-phenylalanine, camphorsulfonic acid, etc., and the bases used are R-phenylethylamine, L-arginine, L-menthol, L-phenylalanine, etc.

[0057] When the compounds mentioned above and below contain phosphorus (P) atoms, the Cahn-Ingold-Prelog rule indicates that the phosphorus atom has a chiral center, and is represented as "R configuration" or "S configuration". If it is not represented as a chiral center, it indicates that the compound is a mixture of "R configuration" and "S configuration".

[0058] The compounds provided by this invention also include isotopic derivatives of formula (I), which refer to one or more atoms in an intermediate or compound being replaced by isotopes of those atoms. The isotopes of formula (I) include, but are not limited to, deuterium (D). 13 C 15 N、 17 O、 18 F, 31Furthermore, the choice of isotope derivatives of the compounds provided by this invention depends on the application field of the compounds. For example, the use of deuterated isotope derivatives can significantly improve the stability of the compound molecules and increase the half-life of the compound molecules in vivo.

[0059] The term "composition" as used above and below can refer to a composition prepared by tightly binding an effective amount of a compound as an active ingredient (including the compound or its solvates, salts, isotope derivatives, etc.) with a pharmaceutically acceptable carrier, diluent, or excipient.

[0060] The term "pharmaceutically acceptable carrier, diluent, or excipient" used above and below can be used in various combinations depending on the desired form of administration. For example, in the preparation of oral dosage forms such as suspensions, syrups, and solutions, diluents such as water, oil, and alcohol can be used; in the preparation of dosage forms such as pills, capsules, powders, or tablets, solid steroids such as starch, sugar, kaolin, lubricants, and binders can be used; and in the preparation of injectables, pH adjusters such as hydrochloric acid, sodium bicarbonate, and phosphate buffer, surfactants such as polyoxyethylene castor oil and lecithin, suspending agents such as gelatin, methylcellulose, and sodium carboxymethylcellulose, and isotonic adjusters such as sodium chloride and glucose can be used.

[0061] The term "therapeutic effective dose" as used above and below refers to an active ingredient administered at approximately 0.01–800 mg / kg, more preferably 0.5–500 mg / kg, at appropriate intervals daily, in a dose form that produces the desired therapeutic effect in the body. Furthermore, the desired therapeutic effect can be achieved by adjusting the dosage based on factors such as disease severity, the age of the recipient, and weight. The "desired therapeutic effect" refers to the ability of the compound or its combinations, administered to the recipient at a therapeutically effective dose, to significantly inhibit tumor growth, induce cancer cell death, reduce tumor size, or decrease or eliminate cancer metastasis and prevent tumor regrowth.

[0062] The compounds of the present invention can be used alone in humans or other mammals to exert anticancer effects and / or in combination with other drugs to enhance the anticancer effect of another anticancer drug.

[0063] The term "halogen" as used above and below refers to the presence of fluorine, chlorine, bromine, or iodine atoms in a molecule.

[0064] The term “heteroatom” as used above and below refers to oxygen, nitrogen, sulfur, boron, or phosphorus atoms contained in a molecule.

[0065] The term "C1-C6 linear or branched alkyl" as used above and below, or in its combination of terms, refers to a straight-chain or branched aliphatic hydrocarbon group containing 1 to 6 carbon atoms, preferably including methyl, ethyl, isopropyl, n-butyl, tert-butyl, etc.; "C1-C16 linear or branched alkyl" as used in its combination of terms refers to a straight-chain or branched aliphatic hydrocarbon group containing 1 to 16 carbon atoms, preferably including n-decyl, 2-methylhexane (isoheptane), 6-methyl-1-octyl, 2-methyldecyl, 2,6-dimethyldecyl, n-undecyl, n-dodecyl, n-tridecyl, n-tetradecyl, n-pentadecanyl, n-hexadecyl, etc.

[0066] The term "C1-C6 linear or branched alkoxy" as used above and below, or in its combination of terms, refers to a group in which one or more "O" atoms or hydroxyl groups are inserted at any reasonable position of "C1-C6 linear or branched alkyl", preferably including methylethoxy, ethylpropoxy, tert-butoxy, ethylisopropoxy, etc.; the term "C1-C16 linear or branched alkoxy" as used in its combination of terms refers to a group in which one or more "O" atoms or hydroxyl groups are inserted at any reasonable position of "C1-C16 linear or branched alkyl", preferably including isopentyl ether, diethylene glycol monohexyl, dipropylene glycol butyl ether, diethylene glycol dibutyl ether, 3,5,5-trimethyl-1-hexyloxy, 10-decoxy, etc.

[0067] The term “C2-C6 linear or branched alkenyl” as used above and below, or in its combined terms, refers to a straight-chain or branched aliphatic hydrocarbon group having 2 to 6 carbon atoms and containing at least one carbon-carbon double bond, preferably including vinyl, propenyl, allyl, n-pentenyl, etc.

[0068] The term “C2-C6 linear or branched alkyne” as used above and below, or in its combined terms, refers to a straight-chain or branched aliphatic hydrocarbon group having 2 to 6 carbon atoms and containing at least one carbon-carbon triple bond, preferably including ethynyl, propynyl, 3-methylbutynyl, pentylyyl, etc.

[0069] The term “C3-C6 cycloalkyl” as used above and below, or in its combination of terms, refers to a cyclic aliphatic hydrocarbon group having 3 to 6 carbon atoms in the molecule, which may be saturated or unsaturated, and may contain one or more carbon-carbon double or triple bonds, preferably including cyclopropyl, cyclopentyl, cyclohexyl, cyclopentenyl, cyclohexenyl, etc.

[0070] The term "C6-C15 aryl" as used above and below, or in its combined terms, refers to a group having 6 to 15 carbon atoms in a molecule that has one or more benzene ring systems, which may be fused or unfused. Fused cyclic structures may be saturated cyclic hydrocarbon groups or unsaturated cyclic hydrocarbon groups, preferably including phenyl, indane, naphthyl, 1,2,3,4-tetrahydronaphthyl, 1,2-dihydroxynaphthyl, etc.

[0071] The terms “(C1-C6 linear or branched alkyl)C6-C15 aryl” or in their combination of terms, as used above and below, which mean “C1-C6 linear or branched alkyl” and “C6-C15 aryl” as defined above, specifically refer to “C6-C15 aryl” groups substituted by straight-chain or branched hydrocarbon groups containing 1 to 6 carbon atoms, preferably including benzyl, phenylpropyl, phenylisopropyl, etc.

[0072] The term "monocyclic or bicyclic heterocyclic alkyl group of 3- to 6-membered rings" as used above and below, or in its combinational terminology, refers to a group containing one or more heteroatoms in a molecule, having a stable 3- to 6-membered cyclic system, each ring having 3 to 6 ring atoms, the group having a single or two cyclic systems, and the cyclic systems may be saturated or unsaturated, preferably including aziridinyl, pyrroleyl, piperazineyl, pyrazineyl, furan, morpholinyl, tetrahydrothiopheneyl, octahydroindolyl, etc.

[0073] The term “5- and / or 6-membered ring monocyclic or bicyclic heteroaryl” as used above and below, or in its combined terminology, refers to a monocyclic or bicyclic group containing one or more heteroatoms and having a stable 5- to 6-membered aromatic ring system, each ring having 5 to 6 ring atoms, preferably including pyrrole, thiophene, pyrazol, isoxazol, pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, 5,6,7,8-tetrahydroquinolinyl, etc.

[0074] The term “C1-C6 linear or branched alkylamino” as used above and below, or in its combination of terms, refers to a group in which a “-N-” or “-NH-” group is inserted at any reasonable position of the “C1-C6 linear or branched alkyl” group, preferably including methylamino, isopropylamino, diisopropylamino, N,N-dimethylbutyl, N-ethylisopropyl, etc. Detailed Implementation Plan Detailed implementation method:

[0076] The compounds of the present invention are synthesized according to the following method:

[0077] Example 1: Preparation of Nu-01

[0078]

[0079] Intermediate M-1

[0080] 2-N-acetylguanine (19.32 g) and N,O-bis(trimethylsilylacetamide) (40.68 g) were added to a dry DCE (600 mL), and the mixture was stirred at 80 °C for 12 hours. After cooling to room temperature, SnCl4 (52.10 g) was added, and stirring continued for 1 hour at room temperature. Tetra-O-acetyl-β-D-ribofuranosyl (31.83 g) was slowly added, and the reaction continued for 24 hours. MeOH (25 mL) was added, and the reaction mixture was diluted with CHCl2 (150 mL), washed with water (500 mL), saturated NaHCO3 / H2O (500 mL), and saturated brine (500 mL), concentrated, and separated by column chromatography to give 23.01 g of intermediate M-1 (yield: 51%) as an off-white solid. MS: ESI 452.4 [M+H] + .

[0081] Intermediate M-2

[0082] Intermediate M-1 (22.56 g) was added to a mixed solution of ammonia (200 mL) and MeOH (200 mL). The mixture was stirred at 60 °C for 24 hours under a closed system. After the reaction was completed, the mixture was concentrated, and the product was recrystallized from acetonitrile / water to give 8.49 g of intermediate M-2 (yield: 60%), which was an off-white solid. MS: ESI 284.1 [M+H] + .

[0083] Intermediate M-3

[0084] 28.32 g of intermediate M-2 and 600 mL of acetonitrile were added to a 1 L three-necked flask. Liquid bromine (32.00 g) was slowly added dropwise to the resulting slurry. The mixture was stirred at room temperature for 6 hours, then quenched with 100 mL of sodium bicarbonate solution (0.01 M). The solution was concentrated under reduced pressure to obtain a pale yellow oily substance. 200 mL of pure water was added and stirred. The mixture was filtered, and the resulting solid was recrystallized from acetonitrile / water to give 23.89 g of intermediate M-3 (yield: 66%), which was an off-white solid. MS: ESI 362.0 [M+H] + .

[0085] Intermediate M-4

[0086] 18.07 g of intermediate M-3 and 1,4-dioxane (400 mL) were added to a 1 L three-necked flask, followed by 1.80 g of [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex, potassium carbonate (8.29 g), and phenylboronic acid (6.10 g). After purging with nitrogen three times, the mixture was heated to reflux for 3 hours. The mixture was filtered, concentrated, and extracted with 300 mL (0.01 M) of hydrochloric acid and 300 mL of ethyl acetate. After concentration, column chromatography was used to separate 10.78 g of intermediate M-4 (yield: 60%) as an off-white solid. MS: ESI 360.2 [M+H] + .

[0087] Intermediate M-5

[0088] 7.18 g of intermediate M-4 and 100 mL of acetone were added to a 250 mL three-necked flask, followed by 5.0 mL of pyridine. 5.17 g of fluorenemethyloxycarbonyl chloride was slowly added at room temperature, and the reaction was allowed to proceed for 12 hours. The system was then concentrated, and 50 mL of ice water was added. A solid precipitated out. After filtration and washing with water, the solid was recrystallized from acetonitrile / water to give 8.02 g of intermediate M-5 (yield: 69%) as a white solid. MS: ESI 582.2 [M+H] + .

[0089] Intermediate M-6

[0090] 5.81 g of intermediate M-5 and 100 mL of acetonitrile were added to a 250 mL three-necked flask, followed by 2.04 g of imidazole and 1.77 g of isopropyl dichlorophosphate. The mixture was reacted at room temperature for 12 hours under nitrogen protection. The system was concentrated, and 25 mL of ice water was added. A solid precipitated out. After filtration and washing with water, the solid was recrystallized from acetonitrile / water to give 4.53 g of intermediate M-6 (yield: 66%) as a white solid. MS: ESI 686.2 [M+H] + .

[0091] Intermediate M-7

[0092] 0.14 g of 1,2,4-triazole was dissolved in 10 mL of anhydrous acetonitrile. 0.30 g of POCl3 was slowly added dropwise under nitrogen protection at 0 °C. After the addition was complete, stirring was continued for 1 hour. 0.55 g of triethylamine was then added dropwise, and stirring continued for another hour. A solution of intermediate M-6 in acetonitrile (1.37 g / 20 mL) was then slowly added dropwise. The reaction was continued at room temperature for 12 hours, followed by concentration under reduced pressure. 10 mL of sodium bicarbonate solution (0.01 M) was added, resulting in the precipitation of a solid. The solid was filtered and recrystallized from acetonitrile / water to give 0.81 g of intermediate M-7 (yield: 55%) as a white solid. MS: ESI 737.4 [M+H] + .

[0093] Nu-01

[0094] Intermediate M-7 (0.74 g) was dissolved in anhydrous acetonitrile (15 mL). 1.0 mL of 50% hydroxylamine aqueous solution was slowly added at 0 °C, and the mixture was stirred at room temperature for 3 hours. 40 mL of ice water was added, and the precipitated solid was added to anhydrous acetonitrile (25 mL). Piperazine (0.40 g) was added, and the mixture was heated to 45 °C and stirred for 2 hours. The mixture was then cooled, concentrated, and separated by column chromatography to obtain 0.14 g of product Nu-01 (yield: 29%), an off-white solid. MS: ESI 479.1 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ:8.28(2H,d,J 6.5),7.54-7.52(3H,m),6.14-6.12(1H,m),4.73-4.71(1H,m),4.66-4.64(1H,m),4.39- 4.37(1H,m),4.25-4.23(1H,m),4.00-3.98(1H,m),3.87-3.85(1H,m),1.26-1.24(6H,m).

[0095] Example 2: Preparation of Nu-02

[0096]

[0097] Nu-02

[0098] The product Nu-01 (4.78 g) was dissolved in anhydrous methanol (150 mL). Ammonia gas was slowly bubbled in at 0–10 °C until the starting material was completely consumed. The bubbling of ammonia gas was then stopped, and the mixture was concentrated under reduced pressure. The residue was recrystallized from acetonitrile / water to give 2.31 g of product Nu-02 (yield: 53%), a white solid. MS: ESI 437.0 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ:8.25(2H,d,J 6.4),7.51-7.49(3H,m),6.13-6.11(1H,m),4.71-4.69(1H,m),4.38-4.36(1H,m),4.20-4.18(1H,m),3.99-3.97(1H,m),3.82-3.80(1H,m).

[0099] Nu-03

[0100] The product Nu-02 (2.18 g) was dissolved in anhydrous acetonitrile (60 mL). Under nitrogen protection, thionyl chloride (0.89 g) was slowly added at 0 °C. After reacting at room temperature for 1 hour, a THF solution of hydroxymethyl isopropyl carbonate (0.67 g) and triethylamine (1.01 g) in 10 mL was slowly added. After the addition was complete, the mixture was reacted at room temperature for 12 hours. The solution was then concentrated under reduced pressure. The residue was separated by column chromatography to obtain 0.52 g of product Nu-03 (yield: 19%), which was an off-white solid. MS: ESI 553.1 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ:8.29(2H,d,J 6.3),7.54-7.51(3H,m),6.29(2H,s),6.15-6.13(1H,m),5.05-5.03(1H,m),4.78-4.7 6(1H,m),4.41-4.39(1H,m),4.21-4.19(1H,m),3.98-3.96(1H,m),1.22-1.20(6H,m).

[0101] Example 3: Preparation of Nu-04

[0102]

[0103] Intermediate M-8

[0104] 35.93 g of intermediate M-4 and 500 mL of acetone were added to a 1000 mL three-necked flask, followed by 20.0 mL of pyridine. Phosgene was slowly bubbled through the mixture at room temperature until the starting material disappeared. The system was concentrated, and 120 mL of ice water was added. A solid precipitated out. The precipitate was filtered, washed with water, and recrystallized from acetonitrile / water to give 33.14 g of intermediate M-8 (yield: 86%), an off-white solid. MS: ESI 386.2 [M+H] + .

[0105] Intermediate M-9

[0106] 19.26 g of intermediate M-8 and 300 mL of acetone were added to a 500 mL three-necked flask, followed by 20.0 mL of pyridine. 12.94 g of fluorenemethyloxycarbonyl chloride was slowly added at room temperature. The reaction was allowed to proceed for 12 hours at room temperature. The system was then concentrated, and 100 mL of ice water was added. A solid precipitated out. After filtration and washing with water, the solid was recrystallized from acetonitrile / water to give 24.30 g of intermediate M-9 (yield: 80%) as an off-white solid. MS: ESI 608.1 [M+H] + .

[0107] M-10

[0108] Intermediate M-9 (60.76 g) was dissolved in DMF (400 mL). POBr3 (28.67 g) was slowly added dropwise under nitrogen protection at 0 °C. After the addition was complete, stirring was continued for 3 hours. Then, 1.2 L of ice water was slowly added, resulting in the precipitation of a large amount of solid. The solid was washed with ice water (200 mL) and dried to obtain 48.28 g of intermediate M-10 (yield: 72%), a pale yellow solid. MS: ESI 670.3 [M+H] + Use it directly in the next step.

[0109] M-11

[0110] Intermediate M-10 (33.52 g) was dissolved in THF (500 mL), and Amberlyst-15 resin (3.56 g) was added. Triethyl phosphite (8.31 g) was slowly added under nitrogen protection. The mixture was heated to 60 °C and reacted for 24 hours. After cooling, the mixture was filtered, concentrated, and the residue was separated by column chromatography to yield 12.00 g of intermediate M-11 (yield: 33%), a pale yellow solid. MS: ESI 728.2 [M+H] + Use it directly in the next step.

[0111] M-12

[0112] 0.69 g of 1,2,4-triazole was dissolved in 100 mL of anhydrous acetonitrile. 2.00 g of POCl3 was slowly added dropwise under nitrogen protection at 0 °C. After the addition was complete, stirring was continued for 1 hour. 3.03 g of triethylamine was then added dropwise, and stirring continued for another hour. A solution of intermediate M-11 in acetonitrile (7.27 g / 100 mL) was then slowly added dropwise. The reaction was continued at room temperature for 12 hours, followed by concentration under reduced pressure. 60 mL of sodium bicarbonate solution (0.01 M) was added, resulting in the precipitation of a solid. The solid was filtered, and recrystallized from acetonitrile / water to give 4.36 g of intermediate M-12 (yield: 56%) as a white solid. MS: ESI 779.2 [M+H] + .

[0113] Nu-04

[0114] Intermediate M-12 (3.89 g) was dissolved in anhydrous acetonitrile (80 mL). 3.0 mL of a 50% hydroxylamine aqueous solution was slowly added at 0 °C, and the mixture was stirred at room temperature for 3 hours. The solution was concentrated, and 30 mL of ice water was added. The precipitated solid was added to methanol (80 mL), followed by barium hydroxide (0.80 g). The mixture was heated to 45 °C and stirred for 3 hours. After cooling, the solution was filtered, concentrated, and separated by column chromatography to obtain 0.52 g of product Nu-04 (yield: 21%), an off-white solid. MS: ESI 495.0 [M+H] + . 1H-NMR(400MHz, CDCl3)δ:8.27(2H,d,J6.3),7.55-7.52(3H,m),6.11-6.10(1H,m),4.70-4.68(1H,m),4.57- 4.55(1H,m),4.21-4.19(5H,m),4.25-4.23(1H,m),1.93-1.91(1H,m),1.72-1.70(1H,m),1.33-1.31(6H,m).

[0115] Example 4: Preparation of Nu-05

[0116]

[0117] M-13

[0118] 4.94 g of product Nu-04 and acetonitrile (80 mL) were added to a 250 mL three-necked flask, followed by the addition of TMSBr (20 mL). The mixture was stirred at room temperature for 48 hours under nitrogen protection. Triethylamine (20 mL) was then added, and the mixture was concentrated to remove acetonitrile. Ice water (10 mL) was added, and the mixture was stirred thoroughly before filtration. The filter cake was directly separated by preparative HPLC, concentrated, and dried to obtain 0.44 g of intermediate M-13 (yield: 10%), a white solid. MS: ESI 439.1 [M+H] + .

[0119] Nu-05

[0120] Intermediate M-13 (0.44 g) was dissolved in anhydrous acetonitrile (50 mL). Under nitrogen protection, thionyl chloride (0.30 g) was slowly added at 0 °C, and the mixture was stirred for 6 hours in an ice bath. Triethylamine (0.44 g) was then slowly added, followed by hydroxymethyl isopropyl carbonate (0.39 g). After the addition was complete, the reaction was stirred for another 48 hours. The mixture was concentrated under reduced pressure at 35 °C, and 0.03 g of product Nu-05 (yield: 4.5%) was obtained directly by preparative liquid chromatography as a white solid. MS: ESI 671.2 [M+H] + . 1 H-NMR(400MHz, CDCl3)δ:8.33(2H,d,J6.5),7.60-7.58(3H,m),6.80-6.78(4H,m),6.13-6.11(1H,m),5.00-4.98(2H,m ),4.79-4.77(1H,m),4.61-4.59(1H,m),4.29-4.27(1H,m),1.93-1.91(2H,m),1.76-1.74(1H,m),1.31-1.29(12H,m).

[0121] Example 5: Preparation of Nu-09

[0122]

[0123] Nu-09

[0124] The product Nu-01 (0.48 g) was dissolved in anhydrous acetonitrile (60 mL). Under nitrogen protection, isobutyryl chloride (0.14 g) and triethylamine (0.30 g) were slowly added at 0 °C. The reaction was carried out at room temperature for 6 hours, and the mixture was concentrated under reduced pressure. The residue was separated by column chromatography to give 0.07 g of product Nu-09 (yield: 13%) as an off-white solid. MS: ESI 549.3 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ:8.33(2H,d,J 6.2),7.56-7.54(3H,m),6.15-6.13(1H,m),4.78-4.76(1H,m),4.69-4.67(1H,m),4.40-4.39(1H,m),4.29- 4.27(1H,m),4.01-3.99(1H,m),3.89-3.88(1H,m),2.66-2.64(1H,m),1.29-1.27(6H,m),1.10-1.07(6H,m).

[0125] Example 6: Preparation of Nu-21

[0126]

[0127] Intermediate M-14

[0128] Uracil (11.21 g) and N,O-bis(trimethylsilylacetamide) (40.68 g) were added to a dry DCE (600 mL), and the mixture was stirred at 80 °C for 12 hours. After cooling to room temperature, SnCl4 (52.10 g) was added, and stirring continued for 1 hour at room temperature. Tetra-O-acetyl-β-D-ribofuranosyl (31.83 g) was slowly added, and the reaction continued for 24 hours. MeOH (25 mL) was added, and the reaction mixture was diluted with CHCl2 (150 mL), washed with water (500 mL), saturated NaHCO3 / H2O (500 mL), and saturated brine (500 mL), concentrated, and separated by column chromatography to give 19.24 g of intermediate M-14 (yield: 52%) as an off-white solid. MS: ESI 371.1 [M+H] + .

[0129] Intermediate M-15

[0130] Intermediate M-14 (18.50 g) was added to a mixed solution of ammonia (200 mL) and MeOH (200 mL). The mixture was stirred at 60 °C for 24 hours under a closed system. After the reaction was completed, the mixture was concentrated, and the product was recrystallized from acetonitrile / water to give 7.81 g of intermediate M-15 (yield: 64%), as a white solid. MS: ESI 245.1 [M+H] + .

[0131] Intermediate M-16

[0132] 2.44 g of intermediate M-15 and 80 mL of acetonitrile were added to a 250 mL three-necked flask, followed by 2.04 g of imidazole and 1.77 g of isopropyl dichlorophosphate. The mixture was reacted at room temperature for 20 hours under nitrogen protection. The system was concentrated, and 50 mL of ice water was added. Extraction was performed with 100 mL of dichloromethane. The organic phase was concentrated, and column chromatography was used to separate 0.77 g of intermediate M-16 (yield: 22%) as a white solid. MS: ESI 349.2 [M+H] + .

[0133] Intermediate M-17

[0134] 0.14 g of 1,2,4-triazole was dissolved in anhydrous acetonitrile (20 mL). 0.40 g of POCl3 was slowly added dropwise under nitrogen protection at 0 °C. After the addition was complete, stirring was continued for 1 hour. Triethylamine (0.95 g) was then added dropwise, and stirring continued for another hour. An acetonitrile solution of intermediate M-16 (0.70 g / 20 mL) was then slowly added dropwise. The reaction was continued at room temperature for 12 hours, followed by concentration under reduced pressure. 20 mL of sodium bicarbonate solution (0.01 M) was added, resulting in the precipitation of a solid. The solid was filtered, and recrystallized from acetonitrile / water to give 0.23 g of intermediate M-17 (yield: 18%), an off-white solid. MS: ESI 400.1 [M+H] + .

[0135] Nu-21

[0136] Intermediate M-17 (0.40 g) was dissolved in anhydrous acetonitrile (15 mL). 1.0 mL of a 50% hydroxylamine aqueous solution was slowly added at 0 °C, and the mixture was stirred at room temperature for 3 hours. 40 mL of ice water was added, and the mixture was extracted with dichloromethane (100 mL). The organic phase was concentrated, and preparative liquid chromatography yielded 0.05 g of product Nu-21 (yield: 15%) as a white solid. MS: ESI 364.1 [M+H] + . 1H-NMR(400MHz, CDCl3)δ:7.07(1H,d,J 6.5),6.01(1H,d,J 6.2),5.71(1H,d,J 5.9),5.39(1H,d,J 7.0),4.61-4.59(1H,m),4.32-4.31(1H,m),4.21-4.19(1H,m),3.97-3.95(1H,m),3.81-3.77(1H,m),1.31-1.29(6H,m).

[0137] Example 7: Nu-22 and its preparation

[0138]

[0139] Nu-22

[0140] The product Nu-21 (3.63 g) was dissolved in anhydrous methanol (150 mL). Ammonia gas was slowly bubbled in at 0–10 °C until the starting material was completely consumed. The ammonia bubbling was then stopped, and the mixture was concentrated under reduced pressure. The residue was separated by preparative liquid chromatography to obtain 0.61 g of product Nu-22 (yield: 19%), which was an off-white solid. MS: ESI 322.0 [M+H] + . 1 H-NMR(400MHz, CDCl3)δ:7.02(1H,d,J 6.3),5.99(1H,d,J 6.1),5.70(1H,d,J 6.9),5.35(1H,d,J 7.2),4.28-4.26(1H,m),4.17-4.15(1H,m),3.93-3.91(1H,m),3.77-3.75(1H,m).

[0141] Nu-23

[0142] The product Nu-22 (0.64 g) was dissolved in anhydrous acetonitrile (60 mL). Under nitrogen protection, thionyl chloride (0.40 g) was slowly added at 0 °C. After reacting in an ice bath for 4 hours, a THF solution of hydroxymethyl isopropyl carbonate (0.34 g) and triethylamine (0.52 g) in 10 mL was slowly added. After the addition was complete, the mixture was reacted in an ice bath for 16 hours. The mixture was then concentrated under reduced pressure. The residue was purified by pre-HPLC to obtain 0.03 g of product Nu-23 (yield: 7%), which was an off-white solid. MS: ESI 438.2 [M+H] + . 1H-NMR(400MHz, CDCl3)δ:7.05(1H,d,J 6.4),6.87(2H,s),6.02(1H,d,J 6.2),5.74(1H,d,J 6.6),5.38(1H,d,J7.1),5.10-5.08(1H,m),4.32-4.30(1H,m),4.19-4.17(1H,m),3.98-3.96(1H,m),3.81-3.79(1H,m),1.26-1.24(6H,m).

[0143] Example 8: Preparation of Nu-37 and Nu-38

[0144]

[0145] Intermediate M-18

[0146] Intermediate M-15 (24.42 g) was added to acetone (300 mL), and sulfuric acid (5 mL) was slowly added dropwise. The mixture was stirred at room temperature for 16 hours. The solution was adjusted to weakly alkaline with sodium bicarbonate (0.01 M), and the acetone was removed by concentration under reduced pressure. The mixture was extracted with ethyl acetate (500 mL), and the organic phase was washed with saturated brine (300 mL). The organic phase was concentrated, and column chromatography was used to separate 21.09 g of intermediate M-18 (yield: 77%) as an off-white solid. MS: ESI 285.1 [M+H] + .

[0147] Intermediate M-19

[0148] Intermediate M-18 (14.20 g) was added to acetonitrile (500 mL), followed by the slow addition of IBX (28.04 g). The mixture was refluxed under nitrogen protection for 6 hours, cooled to room temperature, concentrated under reduced pressure, and extracted with water (400 mL) and dichloromethane (500 mL). The organic phase was washed with saturated brine (300 mL), concentrated, and separated by column chromatography to obtain 4.47 g of intermediate M-19 (yield: 32%) as an off-white solid. MS: ESI 283.0 [M+H] + .

[0149] Intermediate M-20

[0150] Intermediate M-19 (2.80 g) was added to tetrahydrofuran (100 mL), followed by the slow addition of tetramethylmethylene diphosphate (2.74 g) and potassium tert-butoxide (1.60 g). The mixture was refluxed under nitrogen protection for 5 hours, cooled to room temperature, concentrated under reduced pressure, and extracted with water (100 mL). The organic phase was washed with saturated brine (100 mL), concentrated, and separated by column chromatography to obtain 1.13 g of intermediate M-20 (yield: 29%) as an off-white solid. MS: ESI 389.3 [M+H] + .

[0151] Intermediate M-21

[0152] Intermediate M-20 (3.84 g) was added to methanol (100 mL), followed by the slow addition of 10% palladium on carbon (0.40 g). The mixture was refluxed at 0.3 MPa under a hydrogen atmosphere for 15 hours. After cooling to room temperature, the mixture was filtered, concentrated under reduced pressure, and then extracted with water (100 mL). The organic phase was washed with saturated brine (100 mL), concentrated, and separated by column chromatography to obtain 3.01 g of intermediate M-21 (yield: 77%) as an off-white solid. MS: ESI 391.2 [M+H] + .

[0153] Intermediate M-22

[0154] 0.14 g of 1,2,4-triazole was dissolved in anhydrous acetonitrile (20 mL). 0.40 g of POCl3 was slowly added dropwise under nitrogen protection at 0 °C. After the addition was complete, stirring was continued for 1 hour. 0.95 g of triethylamine was then added dropwise, and stirring continued for another hour. A solution of intermediate M-21 in acetonitrile (0.78 g / 20 mL) was then slowly added dropwise. The reaction was continued at room temperature for 12 hours, followed by concentration under reduced pressure. 20 mL of sodium bicarbonate solution (0.01 M) was added, and the mixture was extracted with dichloromethane (30 mL). The organic phase was washed with saturated brine (30 mL), concentrated, and separated by column chromatography to obtain 0.24 g of intermediate M-22 (yield: 27%) as an off-white solid. MS: ESI 442.2 [M+H] + .

[0155] Intermediate M-23

[0156] Intermediate M-22 (0.44 g) was dissolved in anhydrous acetonitrile (25 mL), and 1.0 mL of 50% hydroxylamine aqueous solution was slowly added at 0 °C. The mixture was stirred at room temperature for 5 hours, concentrated under reduced pressure, and liquid-phase separation yielded 0.19 g of intermediate M-23 (yield: 47%) as an off-white solid. MS: ESI 406.1 [M+H] + .

[0157] Nu-37

[0158] Intermediate M-23 (0.20 g) was dissolved in anhydrous acetonitrile (15 mL), and acetic acid (1.0 mL) was slowly added at 0 °C. The mixture was stirred at room temperature for 15 hours under nitrogen protection. Then, 0.01 M sodium bicarbonate solution (15 mL) was added, and the solution was concentrated under reduced pressure. Preparative liquid chromatography yielded 0.09 g of product Nu-37 (yield: 12%) as a white solid. MS: ESI 366.1 [M+H] + . 1 H-NMR(400MHz, CDCl3)δ:7.06(1H,d,J 6.1),5.94(1H,d,J 6.3),5.82-5.80(1H,m),4.49-4.47(1H,m),4.30-4.28(1H,m),3.72-3.7 0(1H,m),3.68(3H,s),3.65(3H,s),1.80-1.78(2H,m),1.65-1.63(2H,m).

[0159] Nu-38

[0160] Nu-37 (0.18 g) was dissolved in anhydrous acetonitrile (10 mL), and TMSBr (0.30 g) was slowly added at 0 °C. The mixture was stirred at room temperature for 30 hours under nitrogen protection. Then, 0.01 M sodium bicarbonate solution (15 mL) was added, and the solution was concentrated under reduced pressure. Liquid chromatography yielded 0.02 g of product Nu-38 (yield: 3%) as a white solid. MS: ESI 338.3 [M+H] + . 1 H-NMR(400MHz, CDCl3)δ:7.02(1H,d,J 6.8),5.91(1H,d,J 6.2),5.79-5.77(1H,m),4.45-4.43(1H,m),4.26-4.24(1H,m),3.70-3.68(1H,m),1.77-1.75(2H,m),1.63-1.61(2H,m).

[0161] Example 9: Preparation of Nu-39

[0162]

[0163] Intermediate M-24

[0164] Nu-38 (0.34 g) was dissolved in anhydrous acetonitrile (25 mL). Isobutyryl chloride (0.43 g) and triethylamine (0.51 g) were slowly added under nitrogen protection at 0 °C. After the addition was complete, the mixture was stirred at room temperature for 15 hours. The solution was concentrated under reduced pressure, and 0.16 g of intermediate M-24 (yield: 29%) was obtained by liquid chromatography as a white solid. MS: ESI 548.0 [M+H] + .

[0165] Nu-39

[0166] Intermediate M-24 (0.27 g) was dissolved in anhydrous acetonitrile (10 mL). Triethylamine (0.20 g) and hexadecyl, 1,3-propylene glycol ether (0.18 g) were slowly added at 0 °C, followed by slow dropwise addition of thionyl chloride (0.09 g). The mixture was stirred at room temperature for 18 hours under nitrogen protection, concentrated under reduced pressure, and the resulting liquid phase was separated to obtain 0.15 g of solid. This solid was dissolved in anhydrous acetonitrile (10 mL), and acetic acid (1.0 mL) was added. The reaction was continued at 0 °C under nitrogen protection for 16 hours. 0.01 M sodium bicarbonate solution (5.0 mL) was added, and the mixture was concentrated under reduced pressure. The resulting liquid phase was separated to obtain 0.03 g of product Nu-39 (yield: 3%), a white solid. MS: ESI 620.1 [M+H] + . 1 H-NMR(400MHz, CDCl3)δ:7.10(1H,d,J 6.9),5.97(1H,d,J 6.3),5.86-5.84(1H,m),4.53-4.51(1H,m),4.33-4.31(1H,m),4.00-3.98(2H,m),3.77-3.75(1H,m),3.40-3 .37(4H,m),2.00-1.98(2H,m),1.80-1.78(2H,m),1.68-1.59(6H,m),1.31-1.25(12H,m),0.95-0.92(2H,m).

[0167] Following the methods described above, and using similar synthetic techniques with commercially available compounds or intermediates, the following compounds were synthesized:

[0168]

[0169]

[0170]

[0171]

[0172] Example 10: Cell viability

[0173] MCF-7 cells (human breast cancer cells) in the logarithmic growth phase were harvested and treated with 1×10⁻⁶ cells. 5 Cells were seeded at a concentration of [number] cells / mL in 96-well plates and incubated at 37°C for 24 hours in a 5% CO2 incubator. Once cells adhered, the compound was prepared to an initial concentration of 20.0 mM (containing 0.02% DMSO for dissolution). The compound solution was then added to the corresponding wells, maintaining a concentration of 20.0 μM per well. The cell zeroing group and blank control group received no compound solution, only an equal volume of culture medium. Each group had three replicates. After adding the compound solution, the cells were incubated at 37°C for another 72 hours in a 5% CO2 incubator. Then, 20.0 μL of MTT solution (5.0 mg / mL) was added, followed by incubation for another 5 hours. The culture medium was removed, and 200 μL of fresh DMSO was added to each well. After shaking thoroughly for 20 minutes, the absorbance at 490 nm was measured for each well. Viability was calculated using the following formula:

[0174] Formula 1: Survival rate % = [1 - (OD)] 490 Drug administration group - OD 490 Blank control group / (OD 490 Cell null group -

[0175] OD 490 (blank control group) × 100%

[0176] The test results are shown in Table 1 below:

[0177] Table 1: Survival rate of compounds on MCF-7 cells

[0178]

[0179]

[0180] The test results showed that the synthesized compounds had a significant inhibitory effect on the proliferation of MCF-7 cells in vitro. In particular, compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 could significantly reduce the survival rate of MCF-7 cells to below 20% in vitro.

[0181] Example 11: In vivo tumor proliferation inhibitory activity of the compound

[0182] An appropriate number of female BALB / c nude mice were used for acclimatization for one week. Cultured human breast cancer MCF-7 cells were then centrifuged with PBS solution and diluted to a concentration of 1×10⁻⁶. 7 Approximately [number] cells / mL, sterilized with ultraviolet light, and inoculated subcutaneously into the right forelimb axilla of nude mice. Mice were fed normally after inoculation until the tumor volume reached 1250±25 mm. 3Successful modeling was considered achieved when the time interval was reached. Sixty-four nude mice weighing 20.0 ± 2.0 g were randomly divided into eight groups of eight mice each: Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, Nu-39, 5-fluorouracil (positive control group), and a blank control group. The compound was dissolved or suspended in a 0.5% sodium carboxymethyl cellulose solution and administered orally at a dose of 30 mg / kg. The blank control group received only an equal volume of 0.5% sodium carboxymethyl cellulose solution. Administration continued for 21 days. During this period, the longest diameter (D) and the maximum transverse diameter (L) of the tumor were measured every 7 days using calipers. The tumor volume was calculated according to Formula 2.

[0183] Formula 2: Mass volume V = D × L 2 (mm 3 )

[0184] The final calculation result is the weighted average of each group as the final tumor volume. The final calculation results are shown in Table 2 below:

[0185] Table 2: Changes in tumor volume in nude mice during drug administration (mean tumor volume ± standard deviation)

[0186]

[0187]

[0188] As shown in Table 2, in the untreated blank control group, the tumor volume increased significantly over time. However, in the Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, Nu-39, and 5-fluorouracil groups, the tumor volume gradually decreased during gavage administration of the compound at a dose of 30 mg / kg. On day 21 of administration, the tumor volume in the treated nude mice was significantly smaller. Compared with the 5-fluorouracil group (positive control group), the tumor volume reduction in the Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 groups was significantly greater, and the average inhibition rate was significantly higher than that in the 5-fluorouracil group. This indicates that the compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 of this invention have a strong inhibitory effect on tumor growth.

[0189] While the above experiments were being conducted, the general condition of the nude mice was recorded. Before successful modeling, all groups of nude mice were active, responsive, had glossy fur, and good appetite. At the end of the experiment, the mice in the blank control group had dull fur, significantly reduced activity, and poor appetite. In contrast, the nude mice in the drug-treated groups were active, agile, had glossy fur, and good appetite, and their general condition was significantly better than that of the nude mice in the 5-fluorouracil group. This indicates that compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 did not exhibit significant side effects due to the toxicity of the compounds in vivo.

[0190] Example 12: Study on the gastric irritation of the compound in mice

[0191] A suitable number of female Kunming mice were acclimatized for one week. Sixty-four mice weighing 20.0 ± 2.0 g were randomly divided into eight groups of eight each: Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, Nu-39, 5-fluorouracil (positive control group), and a blank control group. The compound was dissolved or suspended in 0.5% carboxymethyl cellulose solution and administered by gavage at a dose of 30 mg / kg three times daily. The blank control group received only the corresponding volume of 0.5% carboxymethyl cellulose solution. After 10 consecutive days of administration, the mice were fasted for 24 hours after the last administration on the 10th day, then quickly euthanized by cervical dislocation. The stomach was dissected, cleaned, and soaked in 1.0% formaldehyde solution for 1 hour. The stomach was then opened, and the condition of gastric ulcers was observed and scored according to the criteria in Table 3 below.

[0192] Table 3: Scoring Criteria for Gastric Ulcer Points

[0193] Scoring Criteria Score No or fewer than 5 damage points 0 More than 5 lesions, but no ulcers 1 1-5 obvious ulcer spots 2 Five or more obvious ulcers or one large ulcer 3 Two or more large ulcer spots 4

[0194] The average score of each group of mice was used as the final score, and the results are shown in Table 4 below:

[0195] Table 4: Scores of the mouse gastric irritation test

[0196]

[0197] The scoring results show that although compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 of the present invention exhibit significant local lesions, only a few mice developed gastric ulcers. Compared to 5-fluorouracil, the ulceration was significantly weaker. Therefore, compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 of the present invention do not exhibit significant irritation to the gastric organs compared to 5-fluorouracil.

[0198] Example 13: In vitro cytotoxicity study of the compound HK-2

[0199] HK-2 cells (proximal renal tubular epithelial cell line) in logarithmic growth phase were used at a concentration of 2.0 × 10⁻⁶. 4 Cells were seeded at a density of [number] cells / mL into 96-well plates and incubated at 37°C in a 5% CO2 incubator for 24 hours to allow cell adhesion. Compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, Nu-39, and 5-fluorouracil were prepared to an initial concentration of 20.0 mM (with 0.02% DMSO as a solubilizer). Three replicates were set up for each well. The compound solutions were then added to the corresponding wells, maintaining a final compound concentration of 20.0 μM in each well. The cell nulling group and blank control group received no compound solutions, only an equal volume of culture medium. After addition, the cells were incubated for 48 hours, with the culture medium changed according to standard operating procedures during the experiment. Then add 20.0 μL of MTT solution (5.0 mg / mL), and continue incubation for 5 hours. Remove the culture medium, add 200 μL of fresh DMSO to each well, shake thoroughly for 20 minutes, and then measure the absorbance of each well at 490 nm. Calculate the survival rate using the following formula:

[0200] Formula 3: Survival rate % = [1 - (OD)] 490 Drug administration group - OD 490 Blank control group / (OD 490 Cellular null group - OD 490 (blank control group) × 100%

[0201] The calculation results are shown in Table 5 below:

[0202] Table 5: Survival rate of compounds on HK-2 cells

[0203] compound Survival rate (%) compound Survival rate (%) compound Survival rate (%) Nu-01 89.1±11.1 Nu-29 86.6±9.9 5-Fluorouracil 60.5±7.6 Nu-04 87.2±9.2 Nu-31 87.0±10.2 Nu-25 85.8±13.1 Nu-39 89.4±11.7

[0204] The results showed that the compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 of the present invention had no obvious toxicity to HK-2 cells, which was significantly lower than that of 5-fluorouracil, indicating that the compounds Nu-01, Nu-04, Nu-25, Nu-29, Nu-31, and Nu-39 of the present invention have no obvious nephrotoxicity.

[0205] This application describes several embodiments, but does not limit the invention in any way. There may be more embodiments and implementations within the scope of the embodiments described in this application.

Claims

1. A compound having the structure of formula (I): Or its pharmaceutically acceptable salt, wherein: W is selected from -O-, or does not exist; m can be selected from 1, 2, 3, or 4; R1, R2, and R4 are hydrogen; R3 is selected from hydrogen or hydroxyl groups; G is selected from R 11 Selected from hydrogen, or -(C=O)R5; When W is selected from -O-, L is selected from the form that directly combines with R4 to form a group with the following structural formula (I-1): When W does not exist, L is selected. R5 is selected from C1-C6 linear or branched alkyl, C3-C6 cycloalkyl, and C6-C15 aryl; R7 and R8 are each independently selected from hydrogen, C1-C6 linear or branched alkyl; wherein the alkyl group may be independently and optionally substituted by one or more substituents selected from the following: hydrogen, halogen, C1-C16 linear or branched alkyl, C1-C16 linear or branched alkoxy. R 12 It is phenyl; R 13 Selected from hydrogen, C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, cyano, trifluoromethyl, phenyl, pyridyl; R 14 It is selected from hydrogen, C1-C6 linear or branched alkyl, C1-C6 linear or branched alkoxy, cyano, and trifluoromethyl.

2. The compound according to claim 1, wherein it is a compound of formula (II), The definitions of each generation base in equation (II) are as defined in equation (I) of claim 1.

3. The compound of claim 1, wherein it is a compound of formula (III), The definitions of each substituent in formula (III) are as defined in formula (I) of claim 1.

4. The compound of claim 1, selected from the following compounds:

5. A pharmaceutical composition comprising the compound of any one of claims 1-4, and a pharmaceutical carrier, diluent, or excipient.

6. Use of the compound according to any one of claims 1-4 or the pharmaceutical composition according to claim 5 in the preparation of a medicament for the prevention or treatment of cancer in humans or other mammals.

7. The use according to claim 6, wherein, The cancers mentioned are leukemia, pancreatic cancer, prostate cancer, lung cancer, liver cancer, stomach cancer, colorectal cancer, prostate cancer, bone marrow cancer, thyroid cancer, lymphoma, breast cancer, cervical cancer, or ovarian cancer.

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