New pyrazolopyridinone derivatives and their use in the medical field

Abstract

To address the shortcomings of existing antitumor chemotherapy drugs, this invention provides a class of pyrazolopyridone derivatives and their preparation method. The compounds are structurally designed and modified using pyrazolopyridone as the parent core, resulting in a class of pyrazolopyridone derivatives (I) with good stability and significant antitumor effects: wherein: R 1 The compound is 4-methoxyphenyl,N-phenylbenzamide. This invention relates to the field of tumor treatment, specifically to a class of novel pyrazolopyridone derivatives with advantages such as structural stability and ease of preparation. These compounds can be used to treat diseases such as tumors and condyloma acuminata.

Description

Technical Field

[0001] This invention relates to the field of chemical synthesis of pharmaceuticals, specifically to a novel class of pyrazolopyridone derivatives and their applications. Background Technology

[0002] Cancer, clinically known as malignant tumors, is essentially a pathological state caused by gene mutations in normal cells, resulting in the loss of proliferative regulation. Commonly occurring malignant tumors include lung cancer, cervical cancer, pancreatic cancer, gastric cancer, and liver cancer. The mechanisms of these diseases are highly complex, often resulting from the combined effects of multiple factors, including genetic background, environmental pathogenic factors (such as smoking, ionizing radiation, and exposure to specific chemicals), and viral infections (such as hepatitis B virus and human papillomavirus) (Chinese Journal of Epidemiology, 2024, 45, 621-625). Among the numerous biological characteristics of malignant tumors, invasion and metastasis are the most central and defining features. Cancer cells can penetrate the physiological barrier formed by the basement membrane and extracellular matrix, infiltrating adjacent tissues and further disseminating to distant organs. This not only greatly increases the difficulty of clinical intervention but also seriously threatens the patient's final survival prognosis (Chinese Journal of General Surgery, 2012, 21, 886-892).

[0003] Currently, conventional clinical interventions for cancer mainly rely on surgery, chemotherapy, and radiotherapy. Among these, chemotherapy remains an indispensable cornerstone in the fight against malignant tumors. As a typical representative of classic chemotherapy drugs, cisplatin is widely used in the clinical treatment of various solid tumors due to its excellent tumor-killing efficacy. However, it is undeniable that as a platinum-containing metal complex, cisplatin has revealed many limitations in clinical practice, such as easily inducing significant toxic side effects and easily developing acquired resistance during administration. These shortcomings have largely restricted its wider clinical application (Pharmaceutical Progress, 2020, 44, 243-255). In view of this, exploring and developing lead compounds with novel chemical skeletons and excellent anti-tumor potential has continued to be a core issue in the current field of anti-malignant tumor drug development.

[0004] This invention provides a class of pyrazolopyridone derivatives and their preparation methods. Pyrazolopyridone is a dominant heterocyclic core, possessing three core characteristics: pharmacophore compatibility, excellent drug-likeness, and strong modifiability. The five-membered pyrazolopyridone can form a rigid conjugated plane, with the nitrogen atom and carbonyl group forming multiple hydrogen bond donors / acceptors, easily forming strong hydrogen bonds and π-π stacking with the target site, resulting in strong binding force. 2. Electron isosteric advantage: mimicking ATP adenine, it adapts to the kinase hinge region, exhibiting low nM inhibitory activity and high selectivity. 3. Conformal rigidity + flexible fine-tuning: cyclic fusion restricts conformation, allowing for multiple modification sites. As a dominant core, it has high application value in drug molecule design and optimization. Existing studies have shown that pyrazolopyridone derivatives have broad biological activities and have attracted attention in multiple research directions, including antibacterial, antifungal, antiviral, anti-inflammatory, and antitumor activity. In the field of antitumor drug research and application, compounds containing the pyrazolopyridone skeleton have shown good development potential and have good drug-making potential, providing a useful reference for the design and development of novel antitumor compounds. Summary of the Invention

[0005] To address the shortcomings of existing antitumor chemotherapy drugs, this invention provides a class of pyrazolopyridone derivatives and their preparation method. The compounds are structurally designed and modified using pyrazolopyridone as the parent core, and the new compounds exhibit significant antitumor cell activity.

[0006] This invention relates to a new class of compounds having a pyrazolopyridone structure and their application in the field of tumor treatment.

[0007] The invention is summarized as follows:

[0008] A class of pyrazolopyridone derivatives with good stability and significant antitumor effects (I):

[0009] Where: R 1 It is 4-methoxyphenyl,N-phenylbenzamide.

[0010] The aforementioned class of pyrazolopyridone derivatives (I) is characterized in that such compounds include, but are not limited to: 4-(4-methoxyphenyl)-3-methyl-2,7-dihydro-6H-pyrazolo[3,4-b]pyridin-6-one (I1); 4-(3-methyl-6-oxo-6,7-dihydro-2H-pyrazolo[3,4-b]pyridin-4-yl)-N-benzamide （ I2); Specific preparation scheme The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Furthermore, it should be understood that after reading the teachings of this invention, those skilled in the art can make various alterations or modifications to the invention, and these equivalent forms also fall within the scope defined by the appended claims.

[0011] General solution:

[0012] In step (i), after adding triethylamine to the ethanol mixture of compounds (II) and (III), the reaction solution was heated to reflux at 100°C for 14 hours. After removing the reaction solution by vacuum rotary evaporation, excess n-hexane was added for recrystallization, and the excess filtrate was filtered under reduced pressure to obtain an orange solid (IV). In step (ii), under argon protection, compound (V) was added to the acetic acid solution of compound (IV), and the resulting pale yellow solution was stirred at 160°C for 17 h. After cooling to room temperature, the reaction was detected by TLC to be complete. The reaction solution was then extracted with water and ethyl acetate, and the organic phase was collected and purified by dry column chromatography to obtain the yellowish-white product compound (VI).

[0013] In step (iii), after dissolving compound (VI) in trifluoroacetic acid, the pale yellow reaction solution was refluxed at 160°C for 20 h, and the reaction was confirmed to be complete by TLC. After removing excess trifluoroacetic acid from the reaction solution, an appropriate amount of dichloromethane was added to dissolve the compound, and then an excess of saturated sodium bicarbonate solution was added and stirred overnight to precipitate a white solid. The solid was then filtered to remove excess water, yielding the white solid product compound (I).

[0014] [Example 1] 4-(4-methoxyphenyl)-3-methyl-2,7-dihydro-6H-pyrazolo[3,4-b]pyridin-6-one (I1): In step (i), triethylamine (5.08 ml, 36.54 mmol, 3 equiv) was added to an ethanol mixture of compound (II) (1 g, 12.18 mmol, 1.0 equiv) and compound (III) (1.51 g, 12.18 mmol, 1.0 equiv). The reaction mixture was then refluxed at 100°C for 14 h. After removing the residue by vacuum rotary evaporation, excess n-hexane was added for recrystallization, and the excess filtrate was filtered under reduced pressure to give the orange product (IV) in 64.3% yield. ¹H NMR (400 MHz, DMSO) δ 5.16 (s, 1H), 4.73 (s, 2H), 1.93 (s, 3H), 1.47 (s, 9H). In step (ii), under argon protection, ethyl 3-(4-methoxyphenyl)-3-oxopropionate (3.19 g, 14.36 mmol, 2 equiv) of compound (V1) was added to an acetic acid solution (30 mL) of compound (IV) (1 g, 6.53 mmol, 1 equiv). The resulting pale yellow solution was stirred at 160 °C for 17 h, cooled to room temperature, and the reaction was confirmed to be complete by TLC. The reaction solution was then extracted with a system of water and ethyl acetate, and the organic phase was collected and purified by dry-stirred column chromatography to obtain the yellowish-white product compound (VI1) in 29.9% yield. 1H NMR (400 MHz, DMSO) δ 10.97 (s, 1H), 7.43 (d,J = 8.3 Hz, 2H), 7.06 (d, J = 8.3 Hz, 2H), 6.27 (s, 1H), 3.83 (s, 3H), 2.05(s, 3H), 1.72 (s, 9H). In step (iii), compound (VI1) (100 mg, 321.15 mmol) was dissolved in 5 ml of trifluoroacetic acid. The pale yellow reaction solution was refluxed at 160°C for 20 h, and the reaction was confirmed to be complete by TLC. After removing excess trifluoroacetic acid from the reaction solution, an appropriate amount of dichloromethane was added to dissolve the compound. Then, an excess of saturated sodium bicarbonate solution was added and stirred overnight to precipitate a white solid. The solid product (I1) was obtained by filtration after removing excess water, with a yield of 75.5%. 1H NMR (400 MHz, DMSO) δ12.89 (s, 1H), 11.60 (s, 1H), 7.43 (d, J = 8.4 Hz, 2H), 7.04 (d, J = 8.4 Hz, 2H), 5.85 (s, 1H), 3.82 (s, 3H), 2.06 (s, 3H). 13C NMR (151 MHz, DMSO) δ163.70, 160.19, 148.44, 129.90, 129.63, 114.25, 55.66. HRMS (ESI): m / z calcd.for C14H13N3O2 [M+H]+ 256.1086; found, 256.1087. [Example 2] 4-(3-methyl-6-oxo-6,7-dihydro-2H-pyrazolo[3,4-b]pyridin-4-yl)-N-benzamide (I2): In step (i), triethylamine (5.08 ml, 36.54 mmol, 3 equiv) was added to an ethanol mixture of compound (II) (1 g, 12.18 mmol, 1.0 equiv) and compound (III) (1.51 g, 12.18 mmol, 1.0 equiv). The reaction mixture was then refluxed at 100°C for 14 h. After removing the residue by vacuum rotary evaporation, excess n-hexane was added for recrystallization, and the excess filtrate was filtered under reduced pressure to give the orange product (IV) in 64.3% yield. ¹H NMR (400 MHz, DMSO) δ 5.16 (s, 1H), 4.73 (s, 2H), 1.93 (s, 3H), 1.47 (s, 9H). In step (ii), under argon protection, ethyl (V2)4-(2-ethoxycarbonylacetyl)benzoate (3.45 g, 13.05 mmol, 2 equiv) was added to an acetic acid solution (30 mL) of compound (IV) (1 g, 6.53 mmol, 1 equiv). The resulting pale yellow solution was stirred at 160 °C for 17 h, cooled to room temperature, and after the reaction was confirmed to be complete by TLC, the reaction solution was extracted with water and ethyl acetate, and the organic phase was collected and purified by dry-stirred column chromatography to obtain a yellowish-white product (VI1) in 44.9% yield. 1H NMR (400 MHz, DMSO) δ 11.13 (s, 1H), 8.08 (d, J = 8.1 Hz, 2H), 7.65 (d, J = 8.1 Hz, 2H), 6.36 (s, 1H), 4.36 (q, J = 7.1 Hz, 2H), 1.99 (s,3H), 1.72 (s, 9H), 1.36 (t, J = 7.1 Hz, 3H). In step (iii), compound (VI1) (500 mg, 1.41 mmol) was dissolved in 50 mL of 1 N sodium hydroxide solution and reacted overnight at room temperature. After the reaction was confirmed to be complete by TLC, 4 N dilute hydrochloric acid solution was added to adjust the pH to neutral, and excess water was added to precipitate a large amount of white solid. The solid was then filtered and dried to obtain a white solid product (VII1), with a yield of 65.2%. ¹H NMR (600 MHz, DMSO) δ 13.11 (s, 1H), 11.11 (s, 1H), 8.06 (d, J = 8.2 Hz, 2H), 7.62 (d, J = 8.2 Hz, 2H), 6.35 (s, 1H), 1.99 (s, 3H), 1.73 (s, 9H). In step (IV), compound (VII1) (300 mg, 0.92 mmol) was dissolved in 50 ml of anhydrous ultra-dry DMF, followed by the addition of DIPEA (238.34 mg, 1.84 mmol) and HATU (266.7 mg, 1.11 mmol) and stirring at room temperature for 0.5 h. Then, aniline (103.04 mg, 1.11 mmol) was added and stirred overnight. The reaction was then detected by TLC to indicate that it was complete. After adding excess water and slurrying to precipitate a white solid, the compound (VIII1) was obtained by filtration with a yield of 65.0%. ¹H NMR (600 MHz, DMSO) δ 11.12 (s, 1H), 10.38 (s, 1H), 8.09 (d, J = 7.5 Hz, 2H), 7.82 (d, J = 7.5 Hz, 2H), 7.67 (d, J = 7.5 Hz, 2H), 7.38 (t, J = 7.2 Hz, 2H), 7.12 (t, J = 7.0 Hz, 1H), 6.37 (s, 1H), 2.05 (s, 3H), 1.74 (s, 9H). In step (V), compound (VIII1) (100 mg, 0.250 mmol) was dissolved in 5 ml of trifluoroacetic acid and then refluxed at 160°C. The reaction was confirmed to be complete by TLC. The reaction solution was then slowly poured into a saturated sodium bicarbonate solution under ice bath conditions to adjust the pH to 7 until a yellowish-white solid precipitated. Vacuum filtration was then performed to obtain the product compound (Ⅰ2), with a yield of 81.4%. ¹H NMR (600MHz, DMSO) δ 12.97 (s, 1H), 11.71 (s, 1H), 10.38 (s, 1H), 8.08 (d, J = 8.0Hz, 2H), 7.81 (d, J = 7.9 Hz, 2H), 7.66 (d, J = 8.1 Hz, 2H), 7.37 (t, J = 7.8Hz, 2H), 7.12 (t, J = 7.3 Hz, 1H), 5.95 (s, 1H), 2.05 (s, 3H). ¹³C NMR (151MHz, DMSO) δ 165.48, 139.57, 135.60, 129.10, 128.61, 128.27, 124.23, 120.87.HRMS (ESI): m / z calcd. for C20H16N4O2 [M+H]+ 345.1352; found, 345.1357. [Example 4] In vitro cell proliferation inhibition assay The test cells were human squamous cell carcinoma of the cervix (SiHa).

[0015] The test compound is: 4-(4-methoxyphenyl)-3-methyl-2,7-dihydro-6H-pyrazolo[3,4-b]pyridin-6-one (I1); 4-(3-methyl-6-oxo-6,7-dihydro-2H-pyrazolo[3,4-b]pyridin-4-yl)-N-benzamide （ I2) Log-grown human cervical squamous cells (SiHa) were seeded at a density of 3000 cells per well in two 96-well plates, with a total of 100 µL of cell culture in each well. After 12 h, the cells were allowed to adhere. In one of the 96-well plates, 100 μL of PBS was added after 12 h, followed by 20 μL of MTT solution (5 mg / mL) to each well for incubation for 4 h. The solution in the wells was carefully discarded, and then 150 μL of DMSO was added to each well. After the formazan was completely dissolved, the absorbance at 492 nm was measured using a microplate reader (this value was considered the absorbance at 100% inhibition and was used for subsequent standard curve plotting). Another 96-well plate was incubated with 100 µL of compounds I1 and I2 at concentration gradients of 1000, 500, 250, 125, 62.5, 31.25, 15.625, 7.8125, 3.90625, and 0 nM (total concentration after drug addition) for 48 h. After 48 h, an MTT assay was performed to obtain the absorbance values ​​for each well. Using the absorbance value obtained at 12 h without drug addition as the inhibition rate (100%) and the absorbance value of the blank control group as the inhibition rate (0%), a standard curve was plotted to calculate the inhibition rate for each well. The inhibition rates for each drug-treated well were obtained according to the standard curve. The data were imported into GraphPad Prism software, and the IC50 was calculated. 50 value.

[0016] As shown in Table 1, the experiment showed that the test compound I1 had the most significant anti-proliferative effect on human cervical squamous cell carcinoma cells SiHa. Compared with the broad-spectrum antitumor drug cisplatin, this class of compounds showed more obvious anti-cervical squamous cell carcinoma effects.

[0017] Table 1. Effects of the new compound on the IC50 of SiHa cells. 50 value a

[0018] a These values ​​are the mean ± standard deviation of three replicates.

Claims

1. A class of pyrazolopyridone derivatives, characterized in that... The derivative is a pyrazolopyridone derivative (I) or a pharmaceutically acceptable salt thereof: wherein: In Equation I: R 1 It is 4-methoxyphenyl, N-phenylbenzamide.

2. The pyrazolopyridone derivative (I) according to claim 1, characterized in that... This class of compounds includes, but is not limited to: 4-(4-methoxyphenyl)-3-methyl-2,7-dihydro-6H-pyrazolo[3,4-b]pyridin-6-one (I1), 4-(3-methyl-6-oxo-6,7-dihydro-2H-pyrazolo[3,4-b]pyridin-4-yl)-N-benzamide （ I2).

3. The use of the class of pyrazolopyridone derivatives according to claim 1 and their pharmaceutically acceptable salts, isomers and prodrugs or pharmaceutical compositions in the preparation of diagnoses and treatments for cancer, autoimmune diseases, inflammatory diseases, allergic diseases, allergic reactions, respiratory diseases, cardiovascular diseases, viral infections, transplant rejection, and metabolic / endocrine disorders and neurological disorders.

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