Dihydropyridine small molecule compounds, methods of making and use thereof in the preparation of medicaments for treating neuroblastoma

Abstract

The application relates to a dihydropyridine small-molecule compound and a preparation method and application thereof in preparing a drug for treating neuroblastoma, and belongs to the technical field of drug preparation. The application discloses a dihydropyridine small-molecule compound, a structural formula of which is as shown in the description, which can be used for preparing a drug for treating neuroblastoma with high ALDH18A1 expression, can effectively solve the problems of poor water solubility of YG1702, low inhibition rate of neuroblastoma and other drug application problems, and has a good application prospect.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical preparation technology, and relates to dihydropyridine small molecule compounds, their preparation methods, and their application in the preparation of drugs for treating neuroblastoma. Background Technology

[0002] Neuroblastoma is a neuroendocrine tumor that occurs in the developing sympathetic nervous system. It is the most common extracranial solid tumor in children, with a median age of diagnosis of 18 months. Neuroblastoma exhibits a wide range of clinical behaviors and heterogeneous biological characteristics, from spontaneous regression to progressive disease and metastasis. Current research indicates that neuroblastoma has multiple potential therapeutic targets. For example, the histone-modifying enzyme EZH2 promotes neuroblastoma cell proliferation, and EZH2 inhibitors, in combination with other drugs, can inhibit neuroblastoma growth in vitro. ALK mutations exist in some neuroblastoma tumors, and ALK inhibitors such as crizotinib and lorlatinib can target ALK mutants to inhibit neuroblastoma development. The AKT pathway is an important signaling pathway regulating tumorigenesis and neural differentiation, and the AKT inhibitor Hu7691 has potential differentiation-inducing effects and anti-proliferative activity against various NB cell lines. It interacts with nerves, including inducing neurite growth, cell cycle arrest, and upregulation of neural differentiation markers; inhibitors targeting MDM2, such as Nutlins, SAR405838 (MI77301), and RG7388 (RO5503781 oridasanutlin), can inhibit p53-MDM2 interaction in neuroblastoma and activate p53-induced apoptosis signaling; the Aurora A kinase (AURKA) inhibitor MLN8237 (alisertib) can inhibit neuroblastoma growth in xenograft mouse models by promoting MYCN degradation. Besides small molecule drugs, marketed drugs targeting neuroblastoma also include anti-GD2 antibodies, such as Dinutuximab, which has been approved by the US FDA and included in first-line treatment for neuroblastoma, but has side effects such as severe pain, fever, and low platelet count. In recent years, studies have found that the transmembrane protein GPC2 is a new potential target molecule on the surface of neuroblastoma cells, but clinical evidence regarding the safety and efficacy of GPC2-targeted therapy is still lacking. Currently, several patents protect nucleic acid antibody therapy regimens, such as the application of a transmembrane protein TMEFF1 inhibitor in the preparation of drugs for treating neuroblastoma (CN117582506B) and the application of a TRAF4 inhibitor in the preparation of drugs for treating neuroblastoma (CN202211180257.0). Approximately 20% of patients show MYCN (encoding N-MYC) amplification. As a transforming gene and oncogenic driver, N-MYC plays multiple roles in neuroblastoma malignancies and functions as a major transcriptional regulator, activating genes involved in self-renewal, proliferation, pluripotency, angiogenesis, and metastasis, while inhibiting the expression of genes promoting differentiation, cell cycle arrest, and immune surveillance. Despite the challenges in targeting transcription factors, researchers have attempted to use small or low molecular weight compounds, such as 10058-F4, NY2267, IIA6B17, and Tz-1, which have the ability to target MYC.

[0003] However, these treatments have proven ineffective against neuroblastomas with amplified MYCN. Therefore, there is an urgent need to develop alternative therapeutic strategies targeting MYCN in neuroblastomas. Summary of the Invention

[0004] In view of this, one objective of the present invention is to provide dihydropyridine small molecule compounds; a second objective of the present invention is to provide a method for preparing dihydropyridine small molecule compounds; and a third objective of the present invention is to provide the application of dihydropyridine small molecule compounds in the preparation of drugs for treating neuroblastoma.

[0005] To achieve the above objectives, the present invention provides the following technical solution: 1. A dihydropyridine small molecule compound, the structural formula of which is shown in Formula 1 below: R1 or R2 is selected from -SCH2, H2CO-, -SC(CH3)2, , Any one of them; R3 is selected from , , , , , , , , or Any one of them.

[0006] Preferably, the compound comprises TM-2 to TM-27, and its structural formula is shown below: , , TM-2 TM-3 , , TM-4 TM-5 , TM-6 TM-7 , , TM-8 TM-9 , , TM-10 TM-11 , TM-12 TM-13 , , TM-14 TM-15 , , TM-16 TM-17 , , TM-18 TM-19 , , TM-20 TM-21 , , TM-22 TM-23 , , TM-24 TM-25 , .

[0007] TM-26 TM-27 2. The preparation method of the above-mentioned dihydropyridine small molecule compounds is as follows: o-nitrobenzaldehyde, intermediate compound and NH4HCO3 were added sequentially to an organic solvent under stirring. After sealing, the mixture was reacted in an oil bath at 85-120°C for 2-6 hours. After cooling to room temperature, the mixture was diluted and dissolved with dichloromethane and washed sequentially with saturated sodium bicarbonate and saturated brine. After drying, the mixture was concentrated to obtain a crude product. The crude product was then purified by column chromatography to obtain dihydropyridine small molecule compounds. The intermediate compounds include compounds 8 to 13, and their specific structural formulas are shown below: , , , Compound 8, Compound 9, Compound 10 , , Compound 11, Compound 12, Compound 13 Any one of them.

[0008] Preferably, the organic solvent is ethanol; the preparation method is carried out in a sealed reactor.

[0009] Preferably, the molar ratio of o-nitrobenzaldehyde, the intermediate compound, and NH4HCO3 is 1:1.5 to 4:1 to 2.

[0010] Preferably, the eluent used in the column chromatography separation and purification is a mixture of ethyl acetate and petroleum ether, wherein the volume percentage of ethyl acetate in the mixture is 0-15%.

[0011] 3. Application of the above-mentioned dihydropyridine small molecule compounds in the preparation of drugs for treating neuroblastoma.

[0012] The beneficial effects of this invention are as follows: This invention discloses dihydropyridine small molecule compounds, the structural formula of which is as follows: It can be used in the preparation of drugs for treating neuroblastoma with high ALDH18A1 expression. It can effectively solve the application problems of poor water solubility of YG1702 and low inhibition rate of neuroblastoma. It has good application prospects in the preparation of drugs for treating neuroblastoma.

[0013] Other advantages, objectives, and features of the invention will be set forth in part in the description which follows, and in part will be apparent to those skilled in the art from the following examination, or may be learned from practice of the invention. The objectives and other advantages of the invention can be realized and obtained through the following description. Attached Figure Description

[0014] To make the objectives, technical solutions, and advantages of the present invention clearer, the preferred embodiments of the present invention will be described in detail below with reference to the accompanying drawings, wherein: Figure 1 The dihydropyridine small molecule compound TM-2 in Example 2 1 HNMR spectrum; Figure 2 The mass spectrum of TM-2, a dihydropyridine small molecule compound, is shown in Example 2. Figure 3 The dihydropyridine small molecule compound TM-3 in Example 3 1 HNMR spectrum; Figure 4 The mass spectrum of TM-3, a dihydropyridine small molecule compound, is shown in Example 3. Figure 5 The IC50 of compound YG1702 on SK-N-BE(2) cells 50 Line graph; Figure 6 The IC50 of compound TM-3 on SK-N-BE(2) cells 50 Line graph; Figure 7The IC50 of compound TM-5 on SK-N-BE(2) cells 50 Line graph; Figure 8 The IC50 of compound TM-10 on SK-N-BE(2) cells 50 Line graph; Figure 9 The IC50 of compound TM-12 on SK-N-BE(2) cells 50 Line graph; Figure 10 The IC50 of compound TM-17 on SK-N-BE(2) cells 50 Line graph. Detailed Implementation

[0015] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention. It should be noted that the illustrations provided in the following embodiments are only schematic representations of the basic concept of the present invention. Unless otherwise specified, the following embodiments and features can be combined with each other.

[0016] The structure of YG1702 (TM-1) in the following embodiments is as follows: .

[0017] The dihydropyridine small molecule compounds TM-2 to TM-27 prepared in Examples 2, 3, and 4 are shown below. , , TM-2 TM-3 , , TM-4 TM-5 , TM-6 TM-7 , , TM-8 TM-9 , , TM-10 TM-11 , TM-12 TM-13 , , TM-14 TM-15 , , TM-16 TM-17 , , TM-18 TM-19 , , TM-20 TM-21 , , TM-22 TM-23 , , TM-24 TM-25 , .

[0018] TM-26 TM-27 Example 1 Intermediate compounds 8 to 13 were prepared, and their specific reaction structures are shown below: The specific preparation process is as follows: Trimethyldioxanone (compound 1) (10.0 mmol, 1.0 eq) and any one of compounds 2 to 7 (10.3 mmol, 1.03 eq) are sequentially added to a microwave reaction tube, sealed, and reacted in a microwave reactor at 80–140 °C for 20 minutes. After the reaction is complete, the reaction solution is directly subjected to column chromatography (the eluent is a mixed solution of ethyl acetate and petroleum ether, wherein the percentage of ethyl acetate in the mixture is 0–15%) to obtain a colorless liquid with a yield range of 92–97%.

[0019] Example 2 The specific preparation method for the dihydropyridine small molecule compound TM-2 is shown below: 2.0 mL of ethanol was added to a sealed reactor. Under stirring, o-nitrobenzaldehyde (151 mg, 1.0 mmol, 1.0 eq), intermediate compound 11 (2.5 mmol, 2.5 eq), and NH4HCO3 (103 mg, 1.3 mmol, 1.3 eq) were added sequentially. The reactor was sealed and reacted in an oil bath at 85°C for 4 h. After the reaction was complete, the reaction solution was cooled to room temperature, diluted with 30 mL of dichloromethane, and washed sequentially with saturated sodium bicarbonate and saturated brine. The solution was then dried over anhydrous sodium sulfate and concentrated to obtain a crude product. This crude product was then purified by column chromatography (using a mixture of ethyl acetate and petroleum ether as eluent, with ethyl acetate comprising 40% of the mixture by volume). The purified product yielded 208 mg (0.40 mmol, 40% yield) of a pale yellow slurry, which was identified as the dihydropyridine small molecule compound TM-2. Its structural formula is [structural formula would be inserted here]. .

[0020] Dihydropyridine small molecule compound TM-2 1 H NMR (400 MHz, DMSO) δ 8.98 (s, 1H),7.73 (d, J = 8.1 Hz, 1H), 7.59 (t, J = 7.5 Hz, 1H), 7.46 (d, J = 7.8 Hz, 1H),7.34 (t, J = 7.6 Hz, 1H), 5.61 (s, 1H), 4.13 – 4.04 (m, 2H), 3.95 – 3.87 (m,2H), 3.46 – 3.38 (m, 4H), 3.15 (s, 6H), 2.24 (s, 6H). 13 C NMR (101 MHz, DMSO) δ 166.60, 147.29, 146.39, 142.24, 133.22, 130.86, 127.28, 123.75, 101.39, 69.66, 62.25, 57.93, 34.12, 18.38, [details omitted]. 1 The H NMR spectrum and mass spectrum are shown below. Figure 1 and Figure 2 As shown.

[0021] Example 3 The specific preparation method for the dihydropyridine small molecule compound TM-3 is shown below: 2.0 mL of ethanol was added to a sealed reactor. Under stirring, o-nitrobenzaldehyde (151 mg, 1.0 mmol, 1.0 eq), intermediate compound 8 (510 mg, 2.5 mmol, 2.5 eq), and NH4HCO3 (103 mg, 1.3 mmol, 1.3 eq) were added sequentially. The reactor was sealed and reacted in an oil bath at 85°C for 4 h. After the reaction was complete, the reaction solution was cooled to room temperature, diluted with 30 mL of dichloromethane, and washed sequentially with saturated sodium bicarbonate and saturated brine. The solution was then dried over anhydrous sodium sulfate and concentrated to obtain a crude product. This crude product was then purified by column chromatography (using a mixture of ethyl acetate and petroleum ether as eluent, with ethyl acetate comprising 10% by volume). The purified product yielded 208 mg (0.40 mmol, 40% yield) of a pale yellow slurry, which was identified as the dihydropyridine small molecule compound TM-3, with the following structural formula: .

[0022] Dihydropyridine small molecule compound TM-3 1 The H NMR spectrum and mass spectrum are respectively Figure 3 and Figure 4 As shown, its 1 H NMR(400 MHz, DMSO) δ 9.04 (s, 1H), 7.72 (d, J = 7.8 Hz, 1H), 7.60 (t, J = 7.6Hz, 1H), 7.46 (d, J = 7.3 Hz, 1H), 7.35 (t, J = 7.6 Hz, 1H), 5.56 (s, 1H), 4.09 – 4.02 (m, 2H), 3.93 (dt, J = 11.1, 7.0 Hz, 2H), 2.89 (dt, J = 13.4, 6.7Hz, 2H), 2.61 (t, J = 7.1 Hz, 4H), 2.25 (s, 6H), 1.14 (dd, J = 6.7, 1.9 Hz, 12H). 13 C NMR (101 MHz, DMSO) δ 166.35, 147.29, 146.70, 142.10, 133.32, 130.74,127.40, 123.81, 101.31, 62.60, 34.13, 28.08, 23.28, 18.41.

[0023] Example 4 The specific preparation methods for dihydropyridine small molecule compounds TM-4 to TM-27 are as follows: Dihydropyridine small molecule compound TM-4: Intermediate compound 11 in Example 2 was replaced with intermediate compound 13, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-4. 1 H NMR (400MHz, DMSO) δ 9.00 (s, 1H), 7.50 (ddd, J = 10.9, 9.2, 4.6 Hz, 3H), 7.33 – 7.20(m, 5H), 6.97 – 6.77 (m, 6H), 5.63 (s, 1H), 4.26 (dt, J = 9.2, 4.4 Hz, 2H), 4.10 (ddd, J = 21.9, 9.1, 4.5 Hz, 6H), 2.23 (s, 6H). 13 C NMR (101 MHz, DMSO) δ166.58, 158.17, 147.31, 146.58, 142.07, 133.12, 130.80, 129.47, 127.21,123.75, 120.70, 114.40, 101.28, 65.39, 61.91, 34.21, 18.41.

[0024] Dihydropyridine small molecule compound TM-5: In Example 2, o-nitrobenzaldehyde was replaced with 2-chlorobenzaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-5. 1 H NMR (400 MHz, DMSO) δ 8.92 (s, 1H), 7.33 (dd, J = 7.8,1.6 Hz, 1H), 7.27 – 7.17 (m, 2H), 7.10 (td, J = 7.7, 1.7 Hz, 1H), 5.24 (s,1H), 4.12 – 3.94 (m, 4H), 2.91 (dt, J = 13.4, 6.7 Hz, 2H), 2.71 – 2.54 (m,4H), 2.23 (s, 6H), 1.15 (dd, J = 6.7, 3.6 Hz, 12H). 13C NMR (101 MHz, DMSO) δ166.62, 146.00, 145.93, 131.33, 131.28, 129.01, 127.65, 127.27, 101.47,62.47, 36.88, 34.20, 28.24, 23.29, 18.25.

[0025] Dihydropyridine small molecule compound TM-6: In Example 2, o-nitrobenzaldehyde was replaced with 2-chlorobenzaldehyde, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-6. 1 H NMR (400 MHz, DMSO) δ 8.90 (s, 1H), 7.31 (dd, J = 7.7, 1.5 Hz, 1H), 7.27 – 7.17 (m, 2H), 7.10 (td, J = 7.7, 1.6 Hz, 1H), 5.25 (s, 1H), 4.09 – 3.95 (m, 4H), 3.50 –3.43 (m, 4H), 3.19 (s, 6H), 2.23 (s, 6H). 13 C NMR (101 MHz, DMSO) δ 166.84,145.76, 145.68, 131.44, 131.39, 129.09, 127.59, 127.11, 101.31, 69.82, 62.15,57.98, 37.15, 18.22.

[0026] Dihydropyridine small molecule compound TM-7: In Example 2, o-nitrobenzaldehyde was replaced with 2-hydroxybenzaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-7. 1 H NMR (400 MHz, DMSO) δ 9.72 (s, 1H), 8.86 (s, 1H), 7.38– 7.35 (m, 1H), 7.28 – 7.19 (m, 2H), 7.14 – 7.10 (m, 1H), 5.41 (s, 1H), 4.08– 4.01 (m, 4H), 2.95 (dt, J = 13.2, 6.6 Hz, 2H), 2.75 – 2.68 (m, 4H), 2.28(s, 6H), 1.22 (dd, J = 6.8, 3.6 Hz, 12H). 13C NMR (101 MHz, DMSO) δ 167.22,152.10, 147.86, 136.72, 132.25, 129.83, 128.02, 127.55, 102.13, 63.51, 38.12,34.85, 29.11, 23.67, 19.33.

[0027] Dihydropyridine small molecule compound TM-8: In Example 2, o-nitrobenzaldehyde was replaced with 2-hydroxybenzaldehyde, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-8. 1 H NMR (400MHz, DMSO) δ 9.57 (s, 1H), 8.79 (s, 1H), 7.46 (d, J = 7.8 Hz, 1H), 7.28 –7.20 (m, 2H), 7.16 – 7.12 (m, 1H), 5.32 (s, 1H), 4.05 – 3.97 (m, 4H), 3.62 –3.54 (m, 4H), 3.22 (s, 6H), 2.25 (s, 6H). 13 C NMR (101 MHz, DMSO) δ 166.98,146.22, 145.87, 132.02, 131.66, 129.38, 128.10, 127.53, 102.21, 70.12, 62.65,58.93, 38.26, 19.13.

[0028] Dihydropyridine small molecule compound TM-9: In Example 2, o-nitrobenzaldehyde was replaced with 2-furanaldehyde, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-9. 1 H NMR (400 MHz, DMSO-d6) δ 8.98 (s, 1H), 7.38 (s, 1H), 6.25 (dd, J = 3.2, 1.8 Hz, 1H), 5.88(d, J = 3.2 Hz, 1H), 5.06 (s, 1H), 412 – 4.14 (m, 4H), 3.50 – 3.52 (m, 4H), 3.26 (s, 6H), 2.25 (s, 6H). 13C NMR (101 MHz, DMSO) δ 167.15, 159.03, 147.21, 141.51, 110.64, 104.48, 98.84, 70.50, 62.95, 58.57, 33.17, 18.64.

[0029] Dihydropyridine small molecule compound TM-10: In Example 2, o-nitrobenzaldehyde was replaced with 2-furan carbaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-10. 1 H NMR (400 MHz, DMSO-d6) δ 9.01 (s, 1H), 7.38 (s, 1H), 6.25 (dd, J = 3.2, 1.9 Hz, 1H), 5.89 (d, J = 3.1 Hz, 1H), 5.06 (s, 1H), 4.15(t, J = 6.8 Hz, 4H), 2.97 (hept, J = 6.7 Hz, 2H), 2.73 (t, J = 6.8 Hz, 4H), 2.26 (s, 6H), 1.19 (dd, J = 6.7, 2.3 Hz, 12H). 13 C NMR (101 MHz, DMSO) δ167.02, 158.99, 147.40, 141.58, 110.60, 104.55, 98.78, 63.45, 34.76, 33.08,28.97, 23.78, 18.74.

[0030] Dihydropyridine small molecule compound TM-11: In Example 2, o-nitrobenzaldehyde was replaced with 2-thiophenecaraldehyde, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-11. 1 H NMR (400MHz, DMSO-d6) δ 9.06 (d, J = 2.5 Hz, 1H), 7.18 (dd, J = 3.6, 1.5 Hz, 1H), 6.83 (d, J = 2.6 Hz, 1H), 6.72 (d, J = 4.0 Hz, 1H), 5.20 (d, J = 4.0 Hz, 1H), 4.14 (t, J = 4.0 Hz, 4H), 3.52 (d, J = 4.0 Hz, 4H), 3.25 (s, 6H), 2.27 (s, 6H). 13C NMR (101 MHz, DMSO) δ 167.07, 152.43, 146.67, 126.97, 123.84, 122.98, 101.75, 70.50, 63.01, 58.57, 34.19, 18.63.

[0031] Dihydropyridine small molecule compound TM-12: In Example 2, o-nitrobenzaldehyde was replaced with 2-thiophenecarboxaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-12. 1 H NMR (400 MHz, DMSO) δ 9.09 (s, 1H), 7.19 (dd, J = 5.1,1.2 Hz, 1H), 6.83 (dd, J = 5.1, 3.5 Hz, 1H), 6.71 (d, J = 3.4 Hz, 1H), 5.19(s, 1H), 4.15 (t, J = 6.7 Hz, 4H), 3.02 – 2.92 (m, 2H), 2.73 (t, J = 6.8 Hz, 4H), 2.27 (s, 6H), 1.20 – 1.17 (m, 12H). 13 C NMR (101 MHz, DMSO) δ 166.48,151.75, 146.38, 126.45, 123.44, 122.57, 101.18, 63.01, 34.28, 33.63, 28.49,23.31, 18.25.

[0032] Dihydropyridine small molecule compound TM-13: In Example 2, o-nitrobenzaldehyde was replaced with 4-chloropyridine-3-carboxaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-13. 1 HNMR (400 MHz, DMSO) δ 9.07 (s, 1H), 8.48 (s, 1H), 8.23 ​​(d, J = 5.3 Hz, 1H), 7.35 (d, J = 5.3 Hz, 1H), 5.21 (s, 1H), 4.05 – 4.01(m, 4H), 2.95 – 2.85 (m, 2H), 2.67 – 2.56 (m, 4H), 2.25 (s, 6H), 1.16 – 1.14(m, 12H). 13C NMR (101 MHz, DMSO) δ 166.25, 152.82, 147.80, 146.87, 140.92, 124.47, 99.99, 62.45, 35.83, 34.18, 28.32, 23.24, 18.22.

[0033] Dihydropyridine small molecule compound TM-14: In Example 2, o-nitrobenzaldehyde was replaced with 4-chloropyridine-3-carboxaldehyde, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-14. 1 H NMR(400 MHz, DMSO) δ 9.05 (s, 1H), 8.45 (s, 1H), 8.23 ​​(d, J = 5.3 Hz, 1H), 7.34(d, J = 5.3 Hz, 1H), 5.21 (s, 1H), 4.03 – 4.01 (m, 4H), 3.44 – 3.42 (m, 4H), 3.20 (s, 6H), 2.24 (s, 6H). 13 C NMR (101 MHz, DMSO) δ 166.45, 152.97, 147.73, 146.75, 141.11, 140.47, 124.52, 99.75, 69.75, 62.17, 57.91, 36.17, 18.16.

[0034] Dihydropyridine small molecule compound TM-15: In Example 2, o-nitrobenzaldehyde was replaced with 4-formyl diphenyl sulfide, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-15. Its ¹H NMR (400 MHz, DMSO-d6) δ 8.93 (s, 1H), 7.38 – 7.31 (m, 2H), 7.30 – 7.24 (m, 3H), 7.24 – 7.16 (m, 4H), 4.88 (s, 1H), 4.14 – 4.04 (m, 4H), 2.96-2.89 (m, 2H), 2.69-2.65 (m, 4H), 2.26 (s, 6H), 1.16 (d, J = 8.0Hz, 12H). 13C NMR (101 MHz, DMSO) δ 167.10, 147.95, 146.59, 135.81, 131.93,131.28, 130.70, 129.90, 129.11, 127.57, 101.76, 63.21, 34.74, 28.97, 23.76,18.77.

[0035] Dihydropyridine small molecule compound TM-16: In Example 2, o-nitrobenzaldehyde was replaced with 4-formyl diphenyl sulfide, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-16. 1 HH NMR (400 MHz, DMSO-d6) δ 8.90 (s, 1H), 7.29 – 7.27 (m, 2H), 7.37-7.33 (m, 3H), 7.19 (s, 4H), 4.87 (s, 1H), 4.14 – 4.01 (m, 4H), 3.50 – 3.43 (m, 4H), 3.22 (s, 6H), 2.26 (s, 6H). 13 C NMR (101 MHz, DMSO) δ 167.21, 148.04, 146.45,135.82, 131.88, 131.27, 130.73, 129.90, 129.09, 127.59, 101.82, 70.45, 62.79, 58.49, 18.66.

[0036] Dihydropyridine small molecule compound TM-17: In Example 2, o-nitrobenzaldehyde was replaced with 5-indolecarboxaldehyde, intermediate compound 11 was replaced with intermediate compound 8, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-17. 1 HNMR (400 MHz, DMSO-d6) δ 10.87 (s, 1H), 8.80 (s, 1H),7.30 (d, J = 4.0 Hz, 1H), 7.24 – 7.19 (m, 2H), 6.99 (dd, J = 8.4, 1.7 Hz,1H), 6.30 (s, 1H), 4.95 (s, 1H), 4.06 (t, J = 8.0 Hz, 4H), 2.95 – 2.91 (m,2H), 2.70-2.66 (m, 4H), 2.27 (s, 6H), 1.17 (d, J = 8.0 Hz, 12H). 13C NMR (101MHz, DMSO) δ 167.57, 145.39, 139.27, 135.07, 127.81, 125.47, 121.77, 118.82,111.11, 103.06, 101.36, 63.21, 34.77, 28.98, 23.75, 18.78.

[0037] Dihydropyridine small molecule compound TM-18: In Example 2, o-nitrobenzaldehyde was replaced with 5-indolecarbaldehyde, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-18. 1 H NMR (400MHz, DMSO-d6) δ 10.86 (s, 1H), 8.77 (s, 1H), 7.29 (s, 1H), 7.29 (s, 1H), 7.24– 7.18 (m, 2H), 6.98 (dd, J = 8.4, 1.6 Hz, 1H), 6.31 (s, 1H), 4.95 (s, 1H), 4.05 (s, 4H), 3.48 (d, J = 8 / 0 Hz, 3H), 3.26 (s, 7H), 2.26 (s, 6H). 13 C NMR(101 MHz, DMSO) δ 167.68, 145.22, 139.33, 135.05, 127.79, 125.43, 121.75,118.84, 111.11, 103.12, 101.39, 70.51, 62.69, 58.56, 18.69.

[0038] Dihydropyridine small molecule compound TM-19: In Example 2, o-nitrobenzaldehyde was replaced with p-tert-butylbenzaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-19. 1 H NMR (400 MHz, DMSO-d6) δ 8.88 (s, 1H), 7.21 (d, J =8.0 Hz, 1H), 7.09 (d, J = 8.0 Hz, 1H), 4.86 (s, 1H), 4.09 (t, J = 4.0 Hz,4H), 2.97-291 (m, 2H), 2.70-2.67 (m, 4H), 2.26 (s, 6H), 1.22 – 1.16 (m, 21H). 13C NMR (101 MHz, DMSO) δ 167.31, 148.48, 146.23, 145.25, 127.34, 125.10,102.13, 63.26, 34.75, 34.46, 31.64, 28.96, 23.76, 18.78.

[0039] Dihydropyridine small molecule compound TM-20: In Example 2, o-nitrobenzaldehyde was replaced with p-tert-butylbenzaldehyde, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-20. 1 H NMR (400MHz, DMSO-d6) δ 8.84 (s, 1H), 7.20 (d, J = 8.0 Hz, 2H), 7.09 (d, J = 8.0 Hz, 2H), 4.86 (s, 1H), 4.12-4.03 (m, 4H), 3.50-3.46 (m, 4H), 3.23 (s, 6H), 2.26 (s, 6H), 1.22 (s, 9H). 13 C NMR (101 MHz, DMSO) δ 167.41, 148.44, 146.05,145.36, 127.34, 125.08, 102.22, 70.51, 62.80, 58.54, 38.44, 34.46, 31.65,18.68.

[0040] Dihydropyridine small molecule compound TM-21: In Example 2, o-nitrobenzaldehyde was replaced with 2-methylpropionaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 2, yielding dihydropyridine small molecule compound TM-21 with the following spectral values: δ 8.73 (s, 1H), 4.19 – 4.09 (m, 4H), 3.76 (d, J = 5.4 Hz, 1H), 3.05 – 2.95 (m, 2H), 2.75 (t, J = 6.7 Hz, 4H), 2.23 (s, 6H), 1.50 – 1.42 (m, 1H), 1.21 (dd, J = 6.7, 2.0 Hz, 12H), 0.66 (d, J = 6.8 Hz, 6H); 13C NMR (101 MHz, DMSO) δ 167.66, 146.12, 99.20, 62.64, 37.92, 35.02, 34.29, 28.56, 23.30, 18.21, 18.16.

[0041] Dihydropyridine small molecule compound TM-22: In Example 2, o-nitrobenzaldehyde was replaced with 2-methylpropionaldehyde, intermediate compound 11 was replaced with intermediate compound 9, and the rest of the preparation process was the same as in Example 2, to obtain dihydropyridine small molecule compound TM-22. 1 H NMR (400 MHz, DMSO) δ 8.69 (s, 1H), 4.19 – 4.08 (m,4H), 3.77 (d, J = 5.3 Hz, 1H), 3.53 (t, J = 4.9 Hz, 4H), 3.27 (s, 6H), 2.22(s, 6H), 1.48 – 1.40 (m, 1H), 0.66 (d, J = 6.9 Hz, 6H); 13 C NMR (101 MHz, DMSO) δ 167.79, 145.97, 99.30, 70.08, 62.13, 58.08, 38.03, 35.05, 18.14, 18.05.

[0042] Dihydropyridine small molecule compound TM-23: In Example 2, o-nitrobenzyl was replaced with 4-(2-bromoethoxy)benzaldehyde, and intermediate compound 11 was replaced with intermediate compound 8. The rest of the preparation process was the same as in Example 3, yielding dihydropyridine small molecule compound TM-23. 1 H NMR (400 MHz, DMSO) δ 8.84 (s, 1H), 7.09 (d,J = 8.7 Hz, 2H), 6.78 (d, J = 8.7 Hz, 2H), 4.82 (s, 1H), 4.23 (t, J = 5.4 Hz,2H), 4.07 (t, J = 6.8 Hz, 4H), 3.75 (t, J = 5.4 Hz, 2H), 2.94 (dt, J = 13.3,6.7 Hz, 2H), 2.68 (td, J = 6.7, 1.9 Hz, 4H), 2.25 (s, 6H), 1.18 (d, J = 6.7Hz, 12H); 13 C NMR (101 MHz, DMSO) δ 166.77, 156.14, 145.51, 140.81, 128.38,113.99, 101.84, 67.64, 62.71, 37.72, 34.27, 31.51, 28.49, 23.29, 18.26.

[0043] Dihydropyridine small molecule compound TM-24: In Example 2, o-nitrobenzaldehyde was replaced with 2-chlorobenzaldehyde, and intermediate compound 11 was replaced with intermediate compound 10. The rest of the preparation process was the same as in Example 3, yielding dihydropyridine small molecule compound TM-24. 1 H NMR (400 MHz, DMSO) δ 8.90 (s, 1H), 7.34 – 7.10 (m, 4H), 5.25 (s, 1H), 4.10 – 4.04 (m, 4H), 2.63 – 2.58 (m, 4H), 2.24 (s, 6H), 2.02 (s, 6H); 13C NMR (101 MHz, DMSO) δ 167.10, 146.39, 146.29, 131.74,129.49, 128.07, 127.68, 101.93, 62.12, 37.39, 32.29, 18.69, 15.26.

[0044] Dihydropyridine small molecule compound TM-25: Intermediate compound 11 in Example 2 was replaced with intermediate compound 10, and the rest of the preparation process was the same as in Example 3, to obtain dihydropyridine small molecule compound TM-25. 1 H NMR(400 MHz, DMSO) δ 9.01 (s, 1H), 7.71 (dd, J = 8.1, 1.2 Hz, 1H), 7.61 – 7.57(m, 1H), 7.47 (dd, J = 7.9, 1.3 Hz, 1H), 7.37 – 7.32 (m, 1H), 5.57 (s, 1H), 4.12 – 4.08 (m, 2H), 4.00 – 3.96 (m, 2H), 2.60 – 2.57 (m, 4H), 2.26 (s, 6H), 2.00 (s, 6H); 13 C NMR (101 MHz, DMSO) δ 166.33, 147.28, 146.52, 142.03, 133.20,130.69, 127.31, 123.71, 101.29, 61.74, 33.96, 31.60, 18.33, 14.68.

[0045] Dihydropyridine small molecule compound TM-26: In Example 2, o-nitrobenzaldehyde was replaced with 2-chlorobenzaldehyde, and intermediate compound 11 was replaced with intermediate compound 12. The rest of the preparation process was the same as in Example 3, yielding dihydropyridine small molecule compound TM-26.1 H NMR (400 MHz, DMSO) δ 8.91 (s, 1H), 7.34 – 7.17 (m,13H), 7.10 (td, J = 7.6, 1.7 Hz, 1H), 5.22 (s, 1H), 4.13 – 3.99 (m, 4H), 3.11(t, J = 6.8 Hz, 4H), 2.22 (s, 6H); 13 C NMR (101 MHz, DMSO) δ 166.52, 145.98,145.86, 135.25, 131.30, 129.07, 129.01, 128.32, 127.57, 127.18, 125.95,101.32, 61.31, 36.90, 30.99, 18.21.

[0046] Dihydropyridine small molecule compound TM-27: Intermediate compound 11 in Example 2 was replaced with intermediate compound 12, and the rest of the preparation process was the same as in Example 3, to obtain dihydropyridine small molecule compound TM-27. 1 H NMR(400 MHz, DMSO) δ 9.02 (s, 1H), 7.71 (d, J = 8.1 Hz, 1H), 7.59 (t, J = 7.4Hz, 1H), 7.45 (d, J = 7.6 Hz, 1H), 7.37 – 7.29 (m, 9H), 7.20 – 7.17 (m, 2H), 5.55 (s, 1H), 4.14 – 4.08 (m, 2H), 4.00 – 3.94 (m, 2H), 3.10 (t, J = 6.5 Hz, 4H), 2.23 (s, 6H); 13 C NMR (101 MHz, DMSO) δ 166.30, 146.71, 141.98, 135.20,130.72, 129.07, 128.32, 127.33, 125.95, 123.80, 101.18, 61.55, 33.96, 30.84,18.37.

[0047] The yields of various dihydropyridine small molecule compounds in the above embodiments are shown in Table 1 below: Table 1. Yields of different dihydropyridine small molecule compounds Product Number Yield (mg) molecular weight number of moles yield TM-2 243 434.4 0.56 56% TM-3 208 522.7 0.40 40% TM-4 214 558.6 0.38 38% TM-5 372 512.1 0.73 73% TM-6 223 423.9 0.53 53% TM-7 187 493.7 0.38 38% TM-8 22 405.4 0.05 5% TM-9 330 379.4 0.87 87% TM-10 326 467.6 0.70 70% TM-11 239 395.5 0.60 60% TM-12 210 483.7 0.43 43% TM-13 286 513.1 0.56 56% TM-14 215 424.9 0.51 51% TM-15 417 585.8 0.71 71% TM-16 367 497.6 0.74 74% TM-17 215 516.7 0.42 42% TM-18 137 428.5 0.32 32% TM-19 269 533.8 0.50 50% TM-20 265 445.6 0.59 59% TM-21 258 443.7 0.58 58% TM-22 310 355.4 0.87 87% TM-23 511 600.6 0.85 85% TM-24 106 456.0 0.23 23% TM-25 132 466.6 0.28 28% TM-26 350 580.2 0.60 60% TM-27 349 590.7 0.59 59% Example 5 Aldehyde dehydrogenase 18A1 (ALDH18A1) is a key enzyme in the synthesis of proline from glutamate. It catalyzes the phosphorylation and reduction of glutamate to pyrroline-5-carboxylate (P5C) and plays a crucial role in regulating glutamine metabolism. ALDH18A1, as a component, regulates melanoma cell proliferation through proline biosynthesis. However, the molecular basis of ALDH18A1's influence on tumorigenesis, particularly on MYCN-amplified neoplasms (NBs), remains elusive and warrants further investigation. Previous reports have suggested multiple mechanisms, including various genetic and epigenetic events, in the oncogenesis of high-risk neuroblastomas. MicroRNA (miRNA) dysregulation is a significant component of this pattern, acting through both oncogenic and tumor-suppressive mechanisms. Studies have shown that transcription factor SP1 is a direct target of miR-29b, and a negative correlation between SP1 and miR-29b expression was observed in an acute myeloid leukemia (AML) patient cohort. SP1 interacts with nuclear factor B (NF-κB) and histone deacetylases (HDACs) to further inhibit miR-29b expression, thereby forming a regulatory SP1 / NF-κB / HDAC / miR-29b network that regulates KIT expression in AML. Furthermore, miR-29b is a miRNA that directly targets MYCN, inhibiting endogenous N-MYC protein in the MYCN-amplified human NB cell line Kelly. Therefore, to gain mechanistic insights into the effects of ALDH18A1 activation on NB cell behavior, the genetic downregulation and pharmacological interference of the shell inhibitor YG1702 on ALDH18A1 both attenuate the growth of MYCN-amplified NBs and downregulate MYCN. Moreover, N-MYC mutually regulates ALDH18A1 expression through direct transcriptional activation, establishing a positive feedback loop between ALDH18A1 and MYCN. Therefore, aldehyde dehydrogenase 18A1 (ALDH18A1) is a promising target for inhibiting neuroblastoma, and the efficacy of compound YG1702 validates the antitumor efficacy of this target. Since the water solubility and inhibition rate of YG1702 are not ideal, this invention, based on the structure, introduces hydrophilic groups or enhances the interaction between small molecules and the ALDH18A1 target.

[0048] The CCK8 cytotoxicity assay was used to detect the toxicity of the dihydropyridine small molecule compounds prepared in Examples 2 to 4 of this invention to SK-N-BE(2) cells. The specific experimental steps are as follows: 1) Trypsin digests SK-N-BE(2) cells in a 10 cm dish to prepare a cell suspension and count the cells.

[0049] 2) Inoculate the cell suspension into 96-well plates with 100 μl per well, at a seeding density of 5000 cells / well, and set up 6 replicate wells.

[0050] 3) Place the culture plate in an incubator and incubate overnight (37℃, 5% CO2) until the cells adhere to the plate.

[0051] 4) The next day, by changing the medium, add 8 different concentrations of the drug (100 μM, 50 μM, 25 μM, 12.5 μM, 6.25 μM, 3.125 μM, 1.5625 μM, and 0.78125 μM) to each well of the culture plate, and set up a blank well (the well without cells) and a control well (the well without the drug).

[0052] 5) Place the culture plate in an incubator and let it stand for 48 hours.

[0053] 6) Add 100 μL of serum-free culture medium containing 10% CCK-8 to each well of the culture plate by changing the medium.

[0054] 7) Place the culture plate in an incubator and incubate for 2 hours.

[0055] 8) Measure the absorbance (OD) at 450 nm using an ELISA reader.

[0056] 9) Use GraphPad Prism 8 for data processing to plot the IC50 of each inhibitor on cells. 50 Line graph.

[0057] The maximum half-maximal inhibitory concentrations (IC50) of compounds YG1702 (TM-1), TM-3 in Example 2, and TM-5, TM-10, TM-12, and TM-17 in Example 4 against SK-N-BE (2) cells were tested. The results are shown in Table 2 below. 50 The curves are as follows: Figure 5 , Figure 6 , Figure 7 , Figure 8 , Figure 9 and Figure 10 As shown, the IC of YG1702 (TM-1) 50 The value is 56.32 μM, which is greater than the corresponding values ​​of TM-3, TM-5, TM-10, TM-12, and TM-17, and the IC of TM-17 is also higher. 50 Its value is 20% of that of YG1702(TM-1), which shows great promise for clinical application.

[0058] Table 2. Maximum half-maximal inhibitory concentrations (MCCs) of different compounds on SK-N-BE (2) cells. Target molecule <![CDATA[IC 50 (μM)]]> YG1702 (TM-1) 56.32 TM-3 48.53 TM-5 21.39 TM-10 22.02 TM-12 19.33 TM-17 11.47 The other dihydropyridine small molecule compounds in Example 5 were subjected to CCK8 cytotoxicity assays using the same method. The results showed that their tumor-inhibiting concentration IC50 values ​​were all lower than those of YG1702, indicating that the dihydropyridine small molecule compounds TM-2 to TM-27 prepared in this invention have good application prospects in the preparation of drugs for the treatment of neuroblastoma.

[0059] In summary, this invention discloses dihydropyridine small molecule compounds with the following structural formula: It can be used in the preparation of drugs for treating neuroblastoma with high ALDH18A1 expression, and can effectively solve the problems of poor water solubility and low inhibition rate of neuroblastoma caused by YG1702, and has good application prospects.

[0060] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.

Claims

1. A dihydropyridine small molecule compound, characterized in that, The structural formula of the compound is shown in Formula 1 below: R1 or R2 is selected from -SCH2, H2CO-, -SC(CH3)2, , Any one of them; R3 is selected from , , , , , , , , or Any one of them.

2. The dihydropyridine small molecule compound according to claim 1, characterized in that, The compounds include TM-2 to TM-27, and their structural formulas are shown below: 、 、 TM-2 TM-3 、 、 TM-4 TM-5 、 TM-6 TM-7 、 、 TM-8 TM-9 、 、 TM-10 TM-11 、 TM-12 TM-13 、 、 TM-14 TM-15 、 、 TM-16 TM-17 、 、 TM-18 TM-19 、 、 TM-20 TM-21 、 、 TM-22 TM-23 、 、 TM-24 TM-25 、 。 3. TM-26 TM-27 The method for preparing the dihydropyridine small molecule compound according to any one of claims 1 to 2 is characterized in that, The specific preparation method is as follows: o-nitrobenzaldehyde, intermediate compound and NH4HCO3 were added sequentially to an organic solvent under stirring. After sealing, the mixture was reacted in an oil bath at 85-120°C for 2-6 hours. After cooling to room temperature, the mixture was diluted and dissolved with dichloromethane and washed sequentially with saturated sodium bicarbonate and saturated brine. After drying, the mixture was concentrated to obtain a crude product. The crude product was then purified by column chromatography to obtain dihydropyridine small molecule compounds. The intermediate compounds include compounds 8 to 13, and their specific structural formulas are shown below: 、 、 Compound 8, Compound 9, Compound 10 、 、 Compound 11, Compound 12, Compound 13 Any one of them.

4. The preparation method according to claim 3, characterized in that, The organic solvent is ethanol; the preparation method is carried out in a sealed reactor.

5. The preparation method according to claim 3, characterized in that, The molar ratio of o-nitrobenzaldehyde, intermediate compound and NH4HCO3 is 1:1.5 to 4:1 to 2.

6. The preparation method according to claim 3, characterized in that, The eluent used in the column chromatography separation and purification is a mixture of ethyl acetate and petroleum ether, wherein the volume percentage of ethyl acetate in the mixture is 0-15%.

7. The use of the dihydropyridine small molecule compound according to any one of claims 1 to 2 in the preparation of a medicament for treating neuroblastoma.

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