Pyrimidine derivative as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of biological medicines, and particularly discloses a pyrimidine derivative as well as a preparation method and application thereof. The structure of the pyrimidine derivative is as shown in formula I in the specification. The novel pyrimidine derivative capable of efficiently inhibiting the LSD1 activity is provided, the preparation process is simple and easy to implement, the good effect of resisting colorectal cancer cell proliferation is shown on the animal level, and safe and effective candidate drug molecules are provided for targeted therapy of colorectal cancer.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biological medicine, and particularly relates to a pyrimidine derivative and a preparation method and application thereof. BACKGROUND

[0002] Lysine-specific demethylase 1 (LSD1, also known as KDM1A) as a core enzyme in the epigenetic regulatory network, through catalyzing the demethylation of histone H3K4me1 / 2 and H3K9me1 / 2, dynamically regulates the opening and closing of gene transcription, and plays a key role in cell fate determination, proliferation and differentiation and other biological processes. In recent years, with the in-depth research of tumor epigenetics, the abnormal expression of LSD1 in various malignant tumors and its close association with tumor malignant phenotype have been gradually revealed, making it a potential target for anti-tumor drug research and development.

[0003] In the molecular mechanism of tumor occurrence, the expression disorder of LSD1 is an important driving force for inducing cell malignant transformation. In acute lymphoblastic leukemia (ALL), LSD1 interacts with the oncogenic protein HOX11, inhibits the expression of apoptosis-related genes (such as BIM) by demethylating H3K4me2, and leads to the blockage of lymphocyte apoptosis; in cervical cancer, LSD1 silences the transcription of the tumor suppressor gene PTEN, activates the PI3K / Akt signaling pathway, and promotes the malignant proliferation of cervical epithelial cells. These studies show that the abnormal activation of LSD1 can break the balance between cell proliferation and apoptosis through epigenetic regulation, providing a molecular basis for tumor occurrence.

[0004] LSD1 has a significant regulatory effect on the enhancement of tumor cell proliferation. In a colorectal cancer model, LSD1 continuously drives the RAS / RAF / MEK signaling pathway by activating the promoter activity of the proto-oncogene KRAS, accelerating the cell cycle progression; in thyroid cancer, LSD1 forms a complex with the transcription factor PAX8, up-regulates the expression of the cell cycle protein Cyclin E, and shortens the G1 phase, so that tumor cells rapidly enter the division phase. Clinical studies have confirmed that the expression level of LSD1 is positively correlated with the proliferative activity of tumors, and patients with tumors with high expression of LSD1 have a poor prognosis, suggesting that it can be used as an important indicator for evaluating the malignant degree of tumors and a potential therapeutic target.

[0005] Tumor invasion and metastasis is the main cause of death in patients, and LSD1 plays an important role in this process by regulating the expression of related genes. In prostate cancer, LSD1 up-regulates the expression of EMT-related genes (such as ZEB1) by cooperating with androgen receptor (AR), promoting tumor cells to acquire invasive ability; in lung cancer, LSD1-mediated H3K9me2 demethylation can activate the transcription of chemokine receptors (such as CXCR4), guiding tumor cells to metastasize to specific organs. These findings provide experimental evidence for blocking tumor metastasis by inhibiting LSD1 activity.

[0006] Abnormal expression of LSD1 is also closely related to drug resistance in tumor treatment, which is one of the main challenges in clinical treatment. In breast cancer chemotherapy, LSD1 enhances the resistance of tumor cells to anthracycline drugs by epigenetically regulating the expression of DNA damage repair genes (such as BRCA1); in melanoma, LSD1 regulates the expression of genes related to the MAPK signaling pathway, leading to drug resistance of tumor cells to BRAF inhibitors. In addition, LSD1 can also reduce the therapeutic effect of immune checkpoint inhibitors by regulating the infiltration of immunosuppressive cells in the tumor microenvironment.

[0007] In summary, LSD1 is involved in multiple pathological processes such as tumor occurrence, proliferation, metastasis and drug resistance through complex epigenetic regulation mechanisms, and its functional abnormalities are important molecular characteristics of malignant tumors. Therefore, drug development targeting LSD1 has important theoretical significance and clinical application value, and is expected to provide new effective strategies for the treatment of malignant tumors and improve the prognosis of patients. SUMMARY

[0008] The purpose of the present application is to solve at least one technical problem in the prior art. Based on this, the first aspect of the present application provides a pyrimidine derivative, the structure of which is shown in formula I,

[0009]

[0010] wherein R is selected from hydrogen, methyl, amino, ethoxy; X is selected from carbon, nitrogen, oxygen;

[0011] Ar is any one of

[0012] As a further preferred embodiment, the structure of the above-mentioned pyrimidine derivative is any one of 7a-7l; more preferably, the structure of the above-mentioned pyrimidine derivative is 7l.

[0013]

[0014] The second aspect of the present application also provides a preparation method of the above-mentioned pyrimidine derivative, comprising the following synthesis route:​

[0015]

[0016] The compound 3 is prepared by a palladium-catalyzed coupling reaction from 2,4,5-trichloropyrimidine 1 and 3-chloro-4-cyanobenzoic acid 2; then the intermediate 5 is prepared by reacting with piperazine derivative 4; the compound 5 is cross-coupled with triisopropylsilyl acetylene to obtain the compound 6; and the final product is prepared by a Click reaction of the compound 6 with aryl azide. The synthesis method is simple, raw materials are easy to obtain, and the functional group compatibility is good.

[0017] The third aspect of the present application further provides an LSD1 inhibitor comprising the above-mentioned pyrimidine derivative or a pharmaceutically acceptable salt thereof.

[0018] The fourth aspect of the present application further provides a pharmaceutical composition comprising the above-mentioned pyrimidine derivative or a pharmaceutically acceptable salt thereof. The pharmaceutical composition comprises at least one of gum arabic, sugar syrup, lanolin and starch. The excipient is stable in nature, has no compatibility taboo with the main drug, does not produce side effects, does not affect the curative effect, is not easy to deform, dry, mold, insect-bored at room temperature, is harmless to the human body, has no physiological effect, does not produce chemical or physical action with the main drug, and does not affect the content determination of the main drug.

[0019] The fifth aspect of the present application further provides the use of the above-mentioned pyrimidine derivative in the preparation of a reagent for diagnosing and / or treating a disease characterized by overexpression of LSD1. The disease is preferably colorectal cancer.

[0020] The beneficial effects of the present application are that the present application provides a new pyrimidine derivative with high efficiency in inhibiting the activity of LSD1, the preparation process is simple and easy to operate, and good anti-colorectal cancer cell proliferation effect is shown at the animal level, which provides a safe and effective candidate drug molecule for targeted treatment of colorectal cancer. DETAILED DESCRIPTION

[0021] The concept and technical effects of the present application will be described below in conjunction with examples for a clear and complete description, so as to fully understand the purpose, scheme and effect of the present application. It should be noted that the examples in the present application and the features in the examples can be combined with each other without conflict.

[0022] The following content relates to the synthesis route of pyrimidine derivatives 7a-7l (i.e. formula I) as follows:

[0023]

[0024] The structures of the pyrimidine derivatives 7a-7l are as follows:

[0025]

[0026] The preparation process of compound 3 is as follows:

[0027] Dissolve 2,4,5-trichloropyrimidine 1 (182.0 mg, 1.0 mmol, 1.0 eq) and 3-chloro-4-cyanobenzoic acid 2 (181.0 mg, 1.0 mmol, 1.0 eq) in 1,4-dioxane, under nitrogen protection, and then add 1,1-bis(diphenylphosphino)ferrocene palladium dichloride (73.2 mg, 0.1 mmol, 0.1 eq), sodium carbonate (212.0 mg, 2.0 mmol, 2.0 eq) in sequence, and react overnight at 80°C. After the raw material is completely reacted, quench the reaction by adding saturated brine, extract with ethyl acetate, combine the organic phases, dry over anhydrous sodium sulfate, concentrate, and then purify by column chromatography to obtain compound 3 (215.0 mg, yield 76%). 1 H NMR (400 MHz, Chloroform-d) δ 8.55 (s, 1H), 7.85 (dd, J = 7.5, 2.0 Hz, 1H), 7.79 (d, J = 7.5 Hz, 1H), 7.76 (d, J = 2.0 Hz, 1H). 13 CNMR (100 MHz, Chloroform-d) δ 161.55, 158.35, 155.22, 135.35, 133.18, 132.73, 131.28, 129.51, 123.34, 121.94, 116.12.

[0028] The structures of the corresponding compounds 5a-5e are as follows:

[0029]

[0030] The preparation process of compound 5 is as follows:

[0031] Dissolve compound 3 (1.0 mmol, 1.0 eq) in ethanol, under nitrogen protection, and then add pyridine derivative 4 (1.2 mmol, 1.2 eq), N,N-diisopropylethylamine (387.0 mg, 3.0 mmol, 3.0 eq) in sequence, and reflux to react. After the raw material is completely reacted, quench the reaction by adding saturated sodium bicarbonate, extract with ethyl acetate, combine the organic phases, dry over anhydrous sodium sulfate, concentrate, and then purify by column chromatography to obtain compounds 5a-5e.

[0032] Among them, the characterization results of compound 5a (245.1 mg, yield 65%) are as follows: 1H NMR (400 MHz, Chloroform-d) δ 8.59 (s, 1H), 7.88 - 7.68 (m, 3H), 3.71 - 3.42 (m, 6H), 2.78 (t, J = 5.2 Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.30 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 160.70, 156.02, 155.03, 135.35, 133.18, 132.73, 131.28, 129.51, 121.94, 121.26, 116.12, 59.93, 58.30, 52.43, 44.54.

[0033] Compound 5b (274.1 mg, yield 79%) was characterized by: 1 H NMR (400 MHz, Chloroform-d) δ 8.59 (s, 1H), 7.86 - 7.76 (m, 3H), 3.58 (t, J = 5.2 Hz, 4H), 2.99 (t, J = 5.2 Hz, 4H), 2.60 (s, 3H). 13 C NMR (100 MHz, Chloroform-d) δ 160.70, 156.02, 155.03, 135.35, 133.18, 132.73, 131.28, 129.51, 121.94, 121.26, 116.12, 52.55, 47.21, 44.73.

[0034] Compound 5c (187.0 mg, yield 56%) was characterized by: 1 H NMR (400 MHz, Chloroform-d) δ 8.60 (s, 1H), 7.86 - 7.75 (m, 3H), 3.94 - 3.63 (m, 8H). 13 C NMR (100 MHz, Chloroform-d) δ 160.70, 156.02, 155.03, 135.35, 133.18, 132.73, 131.28, 129.51, 121.94, 121.26, 116.12, 65.67, 46.09.

[0035] Compound 5d (179.3 mg, yield 54%) was characterized by: 1H NMR (400 MHz, Chloroform-d) δ 8.59 (s, 1H), 7.85 - 7.76 (m, 3H), 3.72 (t, J = 5.4 Hz, 4H), 1.70 (p, J = 5.7 Hz, 2H), 1.55 (p, J = 5.6 Hz, 4H). 13 C NMR (100 MHz, Chloroform-d) δ 160.70, 156.02, 155.03, 135.35, 133.18, 132.73, 131.28, 129.51, 121.94, 121.26, 116.12, 45.48, 25.32, 24.65.

[0036] Compound 5e (204.7 mg, yield 59%) was characterized as follows: 1 H NMR (400 MHz, Chloroform-d) δ 8.59 (s, 1H), 7.90 - 7.74 (m, 3H), 3.61 (dt, J = 12.4, 7.1 Hz, 2H), 2.84 (dt, J = 12.4, 7.1 Hz, 2H), 2.64 (p, J = 7.0 Hz, 1H), 2.02 (dq, J = 14.1, 7.1 Hz, 2H), 1.67 (dq, J = 14.1, 7.1 Hz, 2H), 1.07 (s, 2H). 13 C NMR (100 MHz, Chloroform-d) δ 160.70, 156.02, 155.03, 135.35, 133.18, 132.73, 131.28, 129.51, 121.94, 121.26, 116.12, 48.73, 45.07, 35.03.

[0037] The structures of the corresponding compounds 6a-6e are shown below:

[0038]

[0039] The preparation process of compound 6 is as follows:

[0040] Compound 5 (1.0 mmol, 1.0 eq) was dissolved in N,N-dimethylformamide, under nitrogen protection, and then bis(triphenylphosphine)palladium dichloride (72.0 mg, 0.1 mmol, 0.1 eq), cuprous iodide (19.0 mg, 0.1 mmol, 0.1 eq) and N,N-diisopropyl ethylamine (387.0 mg, 3.0 mmol, 3.0 eq) were added successively, and the reaction was carried out at 100°C overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was used for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and then the crude product was dissolved in tetrahydrofuran. Tetrahydrofuran was removed under reduced pressure, and then tetrabutylammonium fluoride (261.5 mg, 1.0 mmol, 1.0 eq) was added for reaction. After the raw material was completely reacted, saturated sodium chloride was added to quench the reaction, and then tetrahydrofuran was removed under reduced pressure. The organic phases were extracted with ethyl acetate, combined, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and then the crude product was purified by column chromatography to obtain compounds 6a-6e.

[0041] The characterization results of compound 6a (161.5 mg, yield 44%) are as follows: 1 H NMR (400 MHz, Chloroform-d) δ 8.67 (s, 1H), 7.95-7.73 (m, 3H), 3.66-3.51 (m, 6H), 3.07 (s, 1H), 2.78 (t, J = 5.2 Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.29 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 161.96, 161.10, 160.53, 137.71, 133.41, 133.21, 130.99, 129.12, 116.12, 110.74, 106.40, 80.42, 76.11, 59.93, 58.30, 52.43, 44.54.

[0042] The characterization results of compound 6b (171.9 mg, yield 51%) are as follows: 1 H NMR (400 MHz, Chloroform-d) δ 8.68 (s, 1H), 7.88-7.78 (m, 3H), 3.58 (t, J = 5.2 Hz, 4H), 3.06 (s, 1H), 2.99 (t, J = 5.2 Hz, 4H), 2.60 (s, 3H). 13C NMR (100 MHz, Chloroform-d) δ 161.96, 161.10, 160.53, 137.71, 133.41, 133.21, 130.99, 129.12, 116.12, 110.74, 106.40, 80.42, 76.11, 52.55, 47.21, 44.73.

[0043] Compound 6c (155.5 mg, yield 48%) was characterized by: 1 H NMR (400 MHz, Chloroform-d) δ 8.68 (s, 1H), 7.93 - 7.71 (m, 3H), 3.91 - 3.69 (m, 8H), 3.05 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 161.96, 161.10, 160.53, 137.71, 133.41, 133.21, 130.99, 129.12, 116.12, 110.74, 106.40, 80.42, 76.11, 65.67, 46.09.

[0044] Compound 6d (135.2 mg, yield 42%) was characterized by: 1 H NMR (400 MHz, Chloroform-d) δ 8.68 (s, 1H), 7.91 - 7.76 (m, 3H), 3.72 (t, J = 5.4 Hz, 4H), 3.03 (s, 1H), 1.70 (p, J = 5.8 Hz, 2H), 1.55 (p, J = 5.6 Hz, 4H). 13 C NMR (100 MHz, Chloroform-d) δ 161.96, 161.10, 160.53, 137.71, 133.41, 133.21, 130.99, 129.12, 116.12, 110.74, 106.40, 80.42, 76.11, 45.48, 25.32, 24.65.

[0045] Compound 6e (175.2 mg, yield 52%) was characterized by: 1H NMR (400 MHz, Chloroform-d) δ 8.67 (s, 1H), 7.89 - 7.78 (m, 3H), 3.60 (dt, J = 12.5, 7.2 Hz, 2H), 3.07 (s, 1H), 2.83 (dt, J = 12.6, 7.1 Hz, 2H), 2.64 (p, J = 7.0 Hz, 1H), 2.01 (dq, J = 14.0, 7.0 Hz, 2H), 1.66 (dq, J = 14.1, 7.1 Hz, 2H), 1.06 (s, 2H). 13 C NMR (100 MHz, Chloroform-d) δ 161.96, 161.10, 160.53, 137.71, 133.41, 133.21, 130.99, 129.12, 116.12, 110.74, 106.40, 80.42, 76.11, 48.73, 45.07, 35.03.

[0046] Example 1

[0047] A pyrimidine derivative, the structure of which is shown as 7a; the preparation method thereof comprises the following process:

[0048] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), azidobenzene (119.1 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropyl alcohol and water, and under nitrogen protection, copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added, and the reaction was carried out at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, extracted with ethyl acetate, the organic phases were combined and dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7a (403.4 mg, yield 83%). 1 H NMR (400 MHz, Chloroform-d) δ 8.67 (s, 1H), 7.89 - 7.78 (m, 3H), 3.60 (dt, J = 12.5, 7.2 Hz, 2H), 3.07 (s, 1H), 2.83 (dt, J = 12.6, 7.1 Hz, 2H), 2.64 (p, J = 7.0 Hz, 1H), 2.01 (dq, J = 14.0, 7.0 Hz, 2H), 1.66 (dq, J = 14.1, 7.1 Hz, 2H), 1.06 (s, 2H). 13C NMR (100 MHz, Chloroform-d) δ 162.79, 157.39, 153.45, 152.42, 137.71, 136.32, 133.41, 133.21, 130.99, 129.73, 129.12, 127.51, 126.50, 125.33, 120.73, 116.12, 110.74, 59.93, 58.30, 52.43, 44.54.

[0049] Example 2

[0050] A pyrimidine derivative, the structure of which is shown as 7b; the preparation method thereof comprises the following process:

[0051] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), 4-azidobenzonitrile (144.1 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropyl alcohol and water, and copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was carried out at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was extracted. The organic phases were combined and dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7b (434.4 mg, yield 85%). 1 H NMR (400 MHz, Chloroform-d) δ 8.75 (s, 1H), 8.24 (s, 1H), 7.94-7.86 (m, 2H), 7.82 (d, J = 2.0 Hz, 1H), 7.64-7.54 (m, 4H), 3.65-3.52 (m, 6H), 2.78 (t, J = 5.2 Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.31 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 157.39, 153.45, 152.42, 137.71, 136.85, 133.41, 133.21, 130.99, 129.12, 128.45, 126.50, 125.33, 122.13, 118.11, 116.12, 110.74, 108.43, 59.93, 58.30, 52.43, 44.54.

[0052] Example 3

[0053] A pyrimidine derivative, the structure of which is shown as 7c; the preparation method thereof comprises the following process:

[0054] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), 1-azido-4-fluorobenzene (137.1 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropyl alcohol and water, and copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was allowed to proceed at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was added for extraction. The organic phases were combined and dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7d (438.5 mg, yield 87%). 1 H NMR (400 MHz, Chloroform-d) δ 8.75 (s, 1H), 8.26 (s, 1H), 7.93-7.85 (m, 2H), 7.82 (d, J = 1.8 Hz, 1H), 7.54-7.47 (m, 2H), 7.44-7.38 (m, 2H), 3.64-3.52 (m, 6H), 2.78 (t, J = 5.2 Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.30 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 157.39, 153.45, 152.42, 137.71, 134.90, 133.41, 133.21, 130.99, 130.15, 129.12, 126.50, 125.33, 122.52, 116.12, 110.74, 59.93, 58.30, 52.43, 44.54.

[0055] Example 4

[0056] A pyrimidine derivative having a structure as shown in 7d; a preparation method thereof includes the following processes:

[0057] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), 1-azido-4-fluorobenzene (137.1 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropyl alcohol and water, and copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was allowed to proceed at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was added for extraction. The organic phases were combined and dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7d (438.5 mg, yield 87%). 1H NMR (400 MHz, Chloroform-d) δ 8.75 (s, 1H), 8.26 (s, 1H), 7.93 - 7.85 (m, 2H), 7.82 (d, J = 1.8 Hz, 1H), 7.29 - 7.21 (m, 2H), 6.92 - 6.82 (m, 2H), 3.63 - 3.52 (m, 6H), 2.78 (t, J = 5.2 Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.30 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 163.36, 162.79, 161.35, 157.39, 153.45, 152.42, 137.71, 133.41, 133.21, 133.08, 133.06, 130.99, 129.12, 126.50, 125.33, 123.24, 123.18, 117.14, 116.98, 116.12, 110.74, 59.93, 58.30, 52.43, 44.54.

[0058] Example 5

[0059] A pyrimidine derivative, the structure of which is shown as 7e; the preparation method thereof comprises the following process:

[0060] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), 1-(azidomethyl)-4-fluorobenzene (151.1 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropyl alcohol and water, copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was carried out at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, extracted with ethyl acetate, the organic phases were combined and dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7e (321.2 mg, yield 62%). 1 H NMR (400 MHz, Chloroform-d) δ 8.71 (s, 1H), 7.84 - 7.75 (m, 3H), 7.57 (s, 1H), 7.27 (ddt, J = 6.9, 5.9, 1.2 Hz, 2H), 7.07 - 6.98 (m, 2H), 5.49 (d, J = 1.2 Hz, 2H), 3.65 - 3.51 (m, 6H), 2.78 (t, J = 5.2

[0061] Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.30 (s, 1H). 13C NMR (100 MHz, Chloroform-d) δ 163.17, 162.79, 161.15, 157.39, 152.42, 148.43, 137.71, 133.41, 133.21, 130.99, 130.54, 130.51, 130.24, 130.18, 129.12, 126.50, 125.81, 116.12, 115.40, 115.24, 110.74, 59.93, 58.30, 53.30, 52.43, 44.54.

[0062] Example 6

[0063] A pyrimidine derivative, the structure of which is shown as 7f; a method for preparing the same comprises the following process:

[0064] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), 1-(azidomethyl)-4-chlorobenzene (167.6 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropyl alcohol and water, and copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was allowed to proceed at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was added for extraction. The organic phases were combined and dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7f (288.4 mg, yield 54%). 1 H NMR (400 MHz, Chloroform-d) δ 8.72 (s, 1H), 7.85-7.72 (m, 3H), 7.57 (s, 1H), 7.34-7.22 (m, 4H), 5.49 (t, J = 1.0 Hz, 2H), 3.64-3.50 (m, 6H), 2.78 (t, J = 5.2 Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.30 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 157.39, 152.42, 148.43, 137.71, 133.49, 133.41, 133.21, 132.43, 130.99, 129.93, 129.12, 129.02, 126.50, 125.81, 116.12, 110.74, 59.93, 58.30, 53.30, 52.43, 44.54.

[0065] Example 7

[0066] A pyrimidine derivative having a structure as shown in 7g; a method for preparing the same comprising the process of:

[0067] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), 4-(azidomethyl)benzonitrile (158.2 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropyl alcohol and water, and copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was allowed to proceed at room temperature overnight. After the completion of the reaction, the reaction was quenched by adding saturated sodium bicarbonate, and extracted with ethyl acetate. The combined organic phase was dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7g (225.8 mg, yield 43%). 1 H NMR (400 MHz, Chloroform-d) δ 8.73 (s, 1H), 7.89-7.83 (m, 2H), 7.81 (d, J = 1.8 Hz, 1H), 7.68 (s, 1H), 7.60-7.55 (m, 2H), 7.46 (dt, J = 7.6, 1.2 Hz, 2H), 5.49 (d, J = 1.2 Hz, 2H), 3.65-3.51 (m, 6H), 2.78 (t, J = 5.2 Hz, 4H), 2.58 (t, J = 7.2 Hz, 2H), 1.30 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 157.39, 152.42, 148.43, 137.71, 136.07, 133.41, 133.21, 132.52, 130.99, 129.50, 129.12, 126.50, 125.81, 119.34, 116.12, 111.23, 110.74, 59.93, 58.30, 53.30, 52.43, 44.54.

[0068] Example 8

[0069] A pyrimidine derivative having a structure as shown in 7h; a method for preparing the same comprising the process of:

[0070] Compound 6a (367.1 mg, 1.0 mmol, 1.0 eq), 1-(azidomethyl)-4-chloro-3- fluorobenzene (185.0 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropanol and water, and copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was allowed to proceed at room temperature overnight. After the starting material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was added for extraction. The combined organic phase was dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7h (215.3 mg, yield 39%). 1 H NMR (400 MHz, Chloroform-d) δ 8.70 (s, 1H), 8.02 (s, 1H), 7.92 - 7.72 (m, 3H), 7.35 (dd, J = 7.5, 5.7 Hz, 1H), 7.12 (ddt, J = 7.5, 2.1, 1.1 Hz, 1H), 6.99 - 6.88 (m, 1H), 5.49 (d, J = 1.2 Hz, 2H), 3.58 (t, J = 5.1 Hz, 6H), 2.78 (t, J = 5.3 Hz, 4H), 2.58 (t, J = 4.8 Hz, 2H), 1.87 (s, 1H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 159.60, 157.59, 157.39, 152.42, 148.43, 137.71, 136.49, 136.43, 133.41, 133.21, 130.99, 129.30, 129.23, 129.12, 126.50, 126.46, 126.44, 125.81, 119.33, 119.17, 117.66, 117.50, 116.12, 110.74, 59.93, 58.30, 52.43, 52.15, 52.12, 44.54.

[0071] Example 9

[0072] A pyrimidine derivative having a structure as shown in 7i; a preparation method thereof includes the following processes:

[0073] Compound 6b (337.1 mg, 1.0 mmol, 1.0 eq), 1-(azidomethyl)-4-chloro-3- fluorobenzene (185.0 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropanol and water, and under nitrogen protection, copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added, and the reaction was carried out at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was used for extraction. The organic phase was combined and dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7i (229.7 mg, yield 44%). 1 H NMR (400 MHz, Chloroform-d) δ 8.71 (s, 1H), 7.84 (d, J = 7.5 Hz, 1H), 7.80 (dd, J = 7.5, 1.8 Hz, 1H), 7.73 (d, J = 2.0 Hz, 1H), 7.62 (s, 1H), 7.48 (dd, J = 7.5, 5.7 Hz, 1H), 7.00 (ddt, J = 7.5, 2.1, 1.2 Hz, 1H), 6.92 (ddt, J = 8.9, 2.1, 1.0 Hz, 1H), 5.49 (d, J = 1.2 Hz, 2H), 3.58 (t, J = 5.2 Hz, 4H), 2.99 (t, J = 5.2 Hz, 4H), 2.60 (s, 3H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 159.60, 157.59, 157.39, 152.42, 148.43, 137.71, 136.49, 136.43, 133.41, 133.21, 130.99, 129.30, 129.23, 129.12, 126.50, 126.46, 126.44, 125.81, 119.33, 119.17, 117.66, 117.50, 116.12, 110.74, 52.55, 52.15, 52.12, 47.21, 44.73.

[0074] Example 10

[0075] A pyrimidine derivative, the structure of which is shown as 7j; the preparation method thereof comprises the following processes:

[0076] Compound 6c (324.1 mg, 1.0 mmol, 1.0 eq), 1-(azidomethyl)-4-chloro-3- fluorobenzene (185.0 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropanol and water, copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was carried out at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was used for extraction. The combined organic phase was dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7j (208.7 mg, yield 41%). 1 H NMR (400 MHz, Chloroform-d) δ 8.71 (s, 1H), 7.86 (d, J = 1.0 Hz, 2H), 7.81 (d, J = 1.3 Hz, 1H), 7.75 (s, 1H), 7.36 (dd, J = 7.5, 5.7 Hz, 1H), 7.12 (dt, J = 7.3, 1.5 Hz, 1H), 6.94 - 6.86 (m, 1H), 5.49 (d, J = 1.4 Hz, 2H), 3.89 - 3.69 (m, 8H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 159.60, 157.59, 157.39, 152.42, 148.43, 137.71, 136.49, 136.43, 133.41, 133.21, 130.99, 129.30, 129.23, 129.12, 126.50, 126.46, 126.44, 125.81, 119.33, 119.17, 117.66, 117.50, 116.12, 110.74, 65.67, 52.15, 52.12, 46.09.

[0077] Example 11

[0078] A pyrimidine derivative, the structure of which is shown as 7k; the preparation method thereof comprises the following process:

[0079] Compound 6d (322.1 mg, 1.0 mmol, 1.0 eq), 1-(azidomethyl)-4-chloro-3- fluorobenzene (185.0 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropanol and water, and under nitrogen protection, copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added, and the reaction was carried out at room temperature overnight. After the raw material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was extracted. The combined organic phase was dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7k (233.2 mg, yield 46%). 1 H NMR (400 MHz, Chloroform-d) δ 8.70 (s, 1H), 8.01 (s, 1H), 7.85 (d, J = 7.5 Hz, 1H), 7.82 - 7.76 (m, 2H), 7.35 (dd, J = 7.5, 5.7 Hz, 1H), 7.16 - 7.09 (m, 1H), 6.94 (ddt, J = 8.9, 2.1, 1.0 Hz, 1H), 5.49 (d, J = 1.4 Hz, 2H), 3.72 (t, J = 5.4 Hz, 4H), 1.70 (p, J = 5.7 Hz, 2H), 1.55 (p, J = 5.6 Hz, 4H). 13 C NMR (100 MHz, Chloroform-d) δ 162.79, 159.60, 157.59, 157.39, 152.42, 148.43, 137.71, 136.49, 136.43, 133.41, 133.21, 130.99, 129.30, 129.23, 129.12, 126.50, 126.46, 126.44, 125.81, 119.33, 119.17, 117.66, 117.50, 116.12, 110.74, 52.15, 52.12, 45.48, 25.32, 24.65.

[0080] Example 12

[0081] A pyrimidine derivative, the structure of which is shown as 7l; the preparation method thereof comprises the following process:

[0082] Compound 6e (337.1 mg, 1.0 mmol, 1.0 eq), 1-(azidomethyl)-4-chloro-3- fluorobenzene (185.0 mg, 1.0 mmol, 1.0 eq) were dissolved in a mixed solvent of isopropanol and water, and copper sulfate pentahydrate (95.7 mg, 0.5 mmol, 0.5 eq) and sodium ascorbate (198.0 mg, 1.0 mmol, 1.0 eq) were sequentially added under nitrogen protection, and the reaction was allowed to proceed at room temperature overnight. After the starting material was completely reacted, saturated sodium bicarbonate was added to quench the reaction, and ethyl acetate was added for extraction. The combined organic phase was dried with anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain compound 7l (167.0 mg, yield 32%). 1 H NMR (400 MHz, Chloroform-d) δ 8.69 (s, 1H), 8.01 (s, 1H), 7.90-7.79 (m, 2H), 7.76 (d, J = 1.8 Hz, 1H), 7.36 (dd, J = 7.5, 5.7 Hz, 1H), 7.10 (ddt, J = 7.6, 2.2, 1.1 Hz, 1H), 6.95 (ddt, J = 8.9, 2.0, 1.0 Hz, 1H), 5.49 (d, J = 1.2 Hz, 2H), 3.63 (dt, J = 12.4, 7.1 Hz, 2H), 2.87 (dt, J = 12.5, 7.0 Hz, 2H), 2.65 (p, J = 7.0 Hz, 1H), 2.02 (dq, J = 13.9, 7.0 Hz, 2H), 1.74-1.60 (m, 2H), 1.08 (s, 2H). 13 CNMR (100 MHz, Chloroform-d) δ 162.79, 159.60, 157.59, 157.39, 152.42, 148.43, 137.71, 136.49, 136.43, 133.41, 133.21, 130.99, 129.30, 129.23, 129.12, 126.50, 126.46, 126.44, 125.81, 119.33, 119.17, 117.66, 117.50, 116.12, 110.74, 52.15, 52.12, 48.73, 45.07, 35.03.

[0083] Compound performance test

[0084] (1) LSD1 kinase inhibition rate test

[0085] LSD1 kinase inhibition activity was monitored by LSD1 Inhibitor Screening Assay Kit (Cat# 700120, Cayman). Briefly, control group: 120 μL of assay reagent, 20 μL of LSD1, 20 μL of HRP, 10 μL of fluorescent substrate and 10 μL of DMSO were added to 3 test wells; blank group: 140 μL of assay reagent, 20 μL of LSD1, 20 μL of HRP, 10 μL of fluorescent substrate and 10 μL of DMSO were added to 3 test wells; experimental group: 120 μL of assay reagent, 20 μL of LSD1, 20 μL of HRP, 10 μL of fluorescent substrate and 10 μL of test drug were added to 3 test wells; 20 μL of peptide was added to the control group and the experimental group and incubated at 37°C for half an hour. The final fluorescence signal (excitation wavelength: 530-540 nm, emission wavelength: 585-595 nm) was obtained using a TECAN SPARK multifunctional microplate reader. The inhibition rate of the compound on enzyme activity was calculated accordingly. The experimental results are shown in Table 1, and compound 7l has good LSD1 inhibition activity.

[0086] Table 1 Determination of LSD1 inhibition activity of compounds

[0087] Compounds IC 50 (nM) 7a 150.2 7b 154.8 7c 310.2 7d 380.6 7e 400.8 7f 270.4 7g 347.3 7h 460.7 7i 320.3 7j 420.5 7k 18.5 71 3.6

[0088] (2) Determination of cell activity of compounds

[0089] The in vitro anti-proliferative activity of compound 7l on various tumor cell lines was evaluated by CCK-8 experiment. The highest concentration was set to 10 μM, 3-fold gradient dilution, the lowest concentration was 0 μM, a total of 10 concentration gradients, which were respectively incubated with various tumor cell lines for 72 h, and the activity of cells was detected by CCK-8. The data were processed using GraphPad Prism 9, and the IC 50 values were calculated. The results are shown in Table 2. Compound 7l has good activity in colorectal cancer cell lines HCT-116 and SW480.

[0090] Table 2 Screening of cell anti-proliferative activity of LSD1 inhibitors

[0091]

[0092] (3) Animal tumor inhibition experiment of compounds

[0093] To evaluate the anti-tumor ability of the active compounds in vivo, animal experiments of two xenograft tumor models of colorectal cancer cells (HCT-116 and SW480) of BALB / c mice were performed. Specifically, first, a mouse subcutaneous tumor-bearing model was constructed to evaluate the anti-tumor effect of compound 7l on BALB / c mice. First, HCT-116 and SW480 were cultured in a 37°C, 5% carbon dioxide incubator. All mice were raised under standard specific pathogen-free (SPF) conditions. 6-8-week-old BALB / c mice were subcutaneously injected with HCT-116 and SW480 cells. The mice were examined for tumor formation every day to determine the development of the tumor. When the tumor grew to a certain size, the mice were randomly divided into a control group and a dosing group, with 5 mice in each group. The dosing group was intraperitoneally injected with the drug, and the dosing amount was 50 mpk. The tumor diameter was measured every day using a vernier caliper, and the body weight of the mice was recorded. The tumor tissue was removed after 21 days. The tumor volume change was used to evaluate the tumor inhibition effect of the compound. As shown in Table 3, compound 7l has excellent in vivo anti-proliferative activity.

[0094] Table 3 In vivo inhibition rate experiment

[0095]

[0096] The above description is only a preferred embodiment of the present application, and the present application is not limited to the above-described embodiments. As long as the same means achieve the technical effects of the present application, they should all belong to the protection scope of the present application. The technical solutions and / or embodiments thereof within the protection scope of the present application can have various modifications and changes.

Claims

1. A pyrimidine derivative, characterized in that, The structure is shown as Formula I, wherein R is selected from hydrogen, methyl, amino, ethoxy; X is selected from carbon, nitrogen, oxygen; Ar is any one of any one of the following:

2. The pyrimidine derivative according to claim 1, characterized in that, The structure is any one of 7a-7l, 3. The pyrimidine derivative according to claim 2, characterized in that, The structure is shown as 7l.

4. A process for the preparation of a pyrimidine derivative according to any one of claims 1 to 3, characterized in that, The synthesis route comprises the following steps:

5. A LSD1 inhibitor, characterized in that, The pharmaceutical composition comprises an excipient.

6. A pharmaceutical composition, characterized by, The excipient is at least one of gum arabic, sugar syrup, lanolin, and starch.

7. The pharmaceutical composition of claim 6, wherein, 9. Use of the pyrimidine derivative according to any one of claims 1 to 3 for the preparation of a reagent for the diagnosis and / or treatment of a disease characterized by overexpression of LSD1.

8. The pharmaceutical composition of claim 7, wherein, The disease is colorectal cancer. ​ 10. Use according to claim 9, characterized in that, ​

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