WO2022032867A1 - Compounds for inhibiting migration of prostate cancer cells - Google Patents

Abstract

Disclosed are compounds for inhibiting the migration of prostate cancer cells. Such compounds have the effect of inhibiting the in vitro migration of PC3 cells, which provides assistance for the development and research of anti-tumor metastasis drugs, provides a theoretical basis for the design of anti-tumor metastasis drugs, and lays a foundation for the development of biological therapies in the future.

Description

A class of compounds used to inhibit the migration of prostate cancer cells technical field

The invention belongs to the technical field of medicine, and relates to a class of compounds for inhibiting the migration of prostate cancer cells.

Background technique

With the increase of global tumor incidence, my country has become a major country in the world of tumor morbidity and mortality. It is estimated that by 2030, 13.2 million people will die from cancer in the world, of which 1/4 will be in China.

About 90% of cancer-related deaths are caused by tumor metastases from the primary tumor, which are often resistant to existing therapies and thus incurable. As one of the important biological characteristics of malignant tumors, tumor metastasis is not only the main cause of death of patients, but also a major challenge for research. Tumor cell invasion is the key link of tumor metastasis, and invasiveness and metastatic ability are the most basic differences between tumor cells and normal cells. Tumor metastasis is a complex, multi-step and multi-gene regulation process. The metastasis of malignant tumor cells is generally considered to include the following steps: 1) tumor cells detach from the primary tumor and adhere to the basement membrane; 2) tumor cells autocrine and induce tumor stromal cells to secrete proteases to degrade the basement membrane, and then pass through the basement membrane The extracellular matrix infiltrates into the surrounding tissue and adheres to the vascular endothelial cells in this part; 3) penetrates the blood vessel wall into the circulatory system, migrates with the blood flow and stays in the new part, and adheres to the vascular endothelial cells in this part; 4) After passing through the vascular wall and extracellular matrix, tumor cells proliferate locally and induce vascular proliferation, and finally form metastases in specific tissues or organs. Therefore, it is of great significance for anti-tumor therapy to understand the process of tumor cells invasive growth and distant metastasis to surrounding tissues, and to take effective blocking measures.

Aiming at the severe situation of tumor metastasis, most of the deaths caused by malignant tumors are the most active field of tumor therapy research in recent years. Tumor molecular targeted therapy is a small molecule targeted therapy that can directly target cancer cells without damaging normal cells as much as possible. It has attracted much attention because of its high efficiency and safety. Exploring the development of molecular targeting compounds based on the inhibition of tumor cell metastasis will bring new opportunities for tumor therapy.

The research results of the present invention clearly influence the migration of PC3 cells in vitro, which will help the development and research of anti-tumor metastasis drugs, provide theoretical basis for the design of anti-tumor metastasis drugs, and lay a foundation for the development of biological therapy in the future.

SUMMARY OF THE INVENTION

The purpose of the present invention is to provide a class of compounds for inhibiting the migration of prostate cancer cells in view of the deficiencies in the prior art. In the present invention, PCa cell line is used, and after adding the compound, the tumor cell migration experiment is performed to observe whether the compound can inhibit the migration of PCa cells.

The purpose of this invention is to realize through the following technical solutions:

In a first aspect, the present invention relates to a class of compounds having the following structural formula:

Wherein, R ₁ is selected from lower branched alkyl, and R ₂ , R ₃ and R ₄ are selected from lower alkyl or H, respectively.

As an embodiment of the present invention, the lower branched chain alkyl group is a 3-8 carbon alkyl group, and the lower alkyl group is a 1-8 carbon alkyl group.

As a specific embodiment of the present invention, the structural formula of the compound includes:

In the second aspect, the present invention relates to the use of a class of compounds of the present invention in the preparation of a medicament for inhibiting the migration of prostate cancer cells.

As an embodiment of the present invention, in the drug for inhibiting the migration of prostate cancer cells, the effective concentration of the compound is 0.5 μM-2.5 μM.

In a third aspect, the present invention relates to a pharmaceutical composition for inhibiting the migration of prostate cancer cells, the pharmaceutical composition using the compound of the present invention as an active ingredient.

As an embodiment of the present invention, the total effective concentration of the compound in the pharmaceutical composition is 0.5 μM-2.5 μM.

As an embodiment of the present invention, the pharmaceutical composition further includes a pharmaceutically acceptable carrier or excipient.

As an embodiment of the present invention, the compounds of the present invention include NJ-78 and NJ-95.

Compared with the prior art, the present invention has the following beneficial effects:

1) The present invention provides a class with a core structure

The compound; the compound can be used to inhibit the migration of prostate cancer cells;

2) The present invention provides a pharmaceutical composition for inhibiting the migration of prostate cancer cells using the compound of the present invention as an active ingredient.

Description of drawings

Other features, objects and advantages of the present invention will become more apparent by reading the detailed description of non-limiting embodiments with reference to the following drawings:

Figure 1 is a schematic diagram of the effect of NJ-78 and NJ-95 on cell viability;

Figure 2 (a) is a graph of the effect of NJ-78 on tumor cell migration, (b) is a statistical graph of the effect of NJ-78 on tumor cell migration;

Figure 3(a) is a graph showing the effect of NJ-95 on tumor cell migration, and (b) is a statistical graph showing the effect of NJ-95 on tumor cell migration.

detailed description

The present invention will be described in detail below with reference to specific embodiments. The following examples will help those skilled in the art to further understand the present invention, but do not limit the present invention in any form. It should be noted that, for those skilled in the art, several changes and improvements can be made without departing from the inventive concept. These all belong to the protection scope of the present invention.

Example 1. Synthesis of compound NJ-78

The synthetic route of compound NJ-78 is shown in the following formula:

Experimental procedures and results for compound NJ-78:

Step 1: Synthesize 2

Sodium hydride (128 mg, 3.2 mmol, 1.2 eq) was dispersed in anhydrous tetrahydrofuran (15 mL), under nitrogen protection, cooled in an ice-water bath, and a solution of compound 1A (445 mg, 2.67 mmol, 1.0 eq) in tetrahydrofuran (2 mL) was added dropwise thereto. , and the reaction was completed at 0°C for 20 minutes. A solution of compound 1 (723 mg, 2.67 mmol, 1.0 eq) in tetrahydrofuran (3 mL) was added dropwise to the reaction solution, and the reaction was carried out at 0° C. for 40 minutes. The reaction was quenched with water and extracted with ethyl acetate. The organic phases were combined, washed with brine, dried over anhydrous sodium sulfate, and spin-dried to obtain a crude product. The crude product was purified by silica gel column (petroleum ether/ethyl acetate=15/1) to obtain the target compound 2 (800 mg, 74%, white solid).

¹ H NMR (400 MHz, CDCl ₃ ) δ 8.69 (s, 1H), 7.45 (q, J=8.4 Hz, 4H), 4.45 (q, J=7.1 Hz, 2H), 4.36 (q, J=7.1 Hz ,2H),2.56(s,3H),1.45(t,J=7.1Hz,3H),1.39(t,J=7.2Hz,3H),1.36(s,9H).

LCMS:(M+H) ⁺ :402.2.

Step 2: Synthesis 3

Compound 2 (0.8 g, 2.26 mmol, 1.0 eq) was dissolved in ethanol (5 mL) and tetrahydrofuran (5 mL), and an aqueous solution (4 mL) of potassium hydroxide (1.3 g, 22.6 mmol, 10 eq) was added thereto, and refluxed at 50°C 2 hours. After the completion of the reaction was detected by TLC and LCMS, the ethanol was removed by rotary evaporation, and the pH value of the aqueous solution was adjusted to 6 with 2N dilute hydrochloric acid. The precipitated solid was filtered, washed with a small amount of water, and dried to obtain the target compound 3 (0.75 g, 96%, white solid).

¹ H NMR (400MHz, DMSO-d ₆ ) δ 8.15(s, 1H), 7.42(d, J=8.3Hz, 2H), 7.37(d, J=8.3Hz, 2H), 2.55(s, 6H) ,1.29(s,9H).

LCMS: (M+H) ⁺ :346.1.

Step 3: Synthesis 4

Compound 3 (0.75 g, 2.17 mmol, 1.0 eq) and DMF (one drop) were dissolved in dry dichloromethane (15 mL), placed in an ice-water bath under nitrogen protection, oxalyl chloride (0.74 mL, 8.7 mmol, 4 eq. ) was added dropwise to the above solution, warmed to room temperature and stirred for 1 hour, spin-dried to remove the solvent and excess oxalyl chloride to obtain a crude aroyl chloride intermediate. This aroyl chloride intermediate was redissolved in dry dichloromethane (15 mL), placed in an ice-water bath under nitrogen protection, and aluminum trichloride (1.73 g, 13 mmol, 6 eq) was added to it in batches, and the reaction was completed at room temperature. overnight. Anhydrous ethanol (2 mL) was added to the reaction solution, and the reaction was carried out at room temperature for 1 hour. After the reaction was detected by LC-MS, dichloromethane (20 mL) and 1N dilute hydrochloric acid (15 mL) were added to separate the layers, and the aqueous phase was extracted with dichloromethane. (2×20 mL), the organic phases were combined, washed with brine (20 mL), dried, suction filtered, the filtrate was concentrated, and purified by column (petroleum ether/ethyl acetate=15/1) to obtain compound 4 (0.35 g, 45%), Pale yellow solid.

¹ HNMR (400 MHz, CDCl ₃ ) δ 9.31 (s, 1H), 8.61 (d, J=2.1 Hz, 1H), 7.77 (dd, J=8.4, 2.2 Hz, 1H), 7.60 (d, J=8.4 Hz, 1H), 4.44(q, J=7.1Hz, 2H), 2.98(s, 3H), 1.45(t, J=7.1Hz, 3H), 1.42(s, 9H).

LCMS: (M+H) ⁺ :356.1.

Step 4: Synthesize 5

Compound 4 (200mg, 0.56mmol, 1.0eq) was dissolved in methanol (5mL) and tetrahydrofuran (5mL), then sodium hydroxide (112mg, 2.81mmol, 5eq) aqueous solution (5mL) was added dropwise to the above solution, 50 ℃ React overnight. The organic phase was spun off, the aqueous phase was extracted with ethyl acetate (10 mL), the pH of the aqueous phase was adjusted to 3 with 2N dilute hydrochloric acid, the precipitated solid was filtered, washed with a small amount of water and dried to obtain compound 5 (90 mg, 50 %, pale yellow solid).

¹ H NMR (400MHz, DMSO-d ₆ ): δ 13.58 (br s, 1H), 8.98 (s, 1H), 8.38 (d, J=2.0Hz, 1H), 7.89 (dd, J=8.5, 2.1Hz ,1H),7.77(d,J=8.5Hz,1H),2.81(s,3H),1.36(s,9H).

LCMS:(M+H) ⁺ :328.1.

HPLC: 99.93%

Step 5: Synthesize 6

5 (10.0g, 30.5mmol, 1.0eq) was added to DCM (150mL), after three to five drops of DMF were added dropwise, the temperature was lowered to 0°C, (COCl) ₂ (10mL, 122.3mmol, 4.0eq) was added dropwise , After reacting at room temperature for 1 hour, the reaction solution was spin-dried, Acetone (80 mL) was added, NaN ₃ (3.0 g, 46.1 mmol, 1.55 eq) was added, 40 mL of water was added, and the reaction was performed by heating at 70 °C in an oil bath overnight, and detected by LC-MS. The reaction was completed, cooled to room temperature, spin-dried excess acetone, extracted with water and DCM, spin-dried the organic phase, dried and concentrated by column chromatography (50% EtOAc in petroleum ether) and purified to obtain compound 6 (3.5 g, 38%).

¹ H NMR (400MHz, CDCl ₃ ): δ 8.53 (s, 1H), 7.97 (s, 1H), 7.63 (d, J=8.4Hz, 1H), 7.50 (d, J=8.5Hz, 1H), 3.80(brs, 2H), 2.49(s, 3H), 1.33(s, 9H).

LCMS: 299.1([M+H] ⁺ ).

Step 6: Synthesize 7

Compound 6 (3.5 g, 11.7 mmol, 1.0 eq) was dissolved in MeCN (100 mL), to which was added CuI (2.6 g, 14.0 mmol, 1.2 eq) and t-BuONO (4.8 g, 46.9 mmol, 4 eq), 60 °C reaction overnight. LC-MS detected that the reaction was complete. After the acetonitrile was spun out, brine and ammonia water were mixed and added to the reaction system. Then DCM was added for extraction. (2.5 g, 52%).

¹ H NMR (400 MHz, CDCl ₃ ) δ 9.05 (s, 1H), 8.52 (d, J=2.2 Hz, 1H), 7.68 (dd, J=8.5, 2.2 Hz, 1H), 7.52 (d, J= 8.5Hz, 1H), 2.78(s, 3H), 1.34(s, 9H).

LCMS: 410.2([M+H] ⁺ ).

Step 7: Synthesize 8

Compound 7 (500 mg, 1.2 mmol, 1.0 eq) was dissolved in dry dioxane (10 mL), to which was added B ₂ Pin ₂ (1.9 g, 7.2 mmol, 6.0 eq), KOAc (239 mg, 2.4 mmol, 2.0 eq) and Pd(dppf)Cl ₂ (89 mg, 0.12 mmol, 0.1 eq), and reacted at 90° C. for 15 hours. LC-MS detected the reaction was complete, after cooling, concentrated to obtain crude product. Extract with water and dichloromethane (50 mL x 3). The combined organic phases were washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, and spin-dried to obtain the crude compound. The crude compound was purified by silica gel column (petroleum ether/EtOAc=100:1) to give crude compound 8 (400 mg) as a white solid.

¹ H NMR (400 MHz, CDCl ₃ ) δ 9.07 (s, 1H), 8.54 (d, J=2.2 Hz, 1H), 7.65 (dd, J=8.5, 2.3 Hz, 1H), 7.50 (d, J= 8.4Hz, 1H), 2.78(s, 3H), 1.34(s, 9H), 1.31(s, 12H).

LCMS: 410.2([M+H] ⁺ ).

Step 8: Synthesize 9

Compound 8 (1.9 g, crude) was added to THF (30 mL) and H ₂ O (10 mL) to dissolve, then NaIO ₄ (6.0 g, 4.8 mmol, 5 eq) was added, the reaction was carried out at 50° C. for 15 h, THF was spin-dried, and DCM was used. Extracted three times, washed with brine again, the organic phase was dried over anhydrous Na ₂ SO ₄ , and spin-dried to obtain crude compound 9 (280 mg) as a yellow solid.

LCMS: (M+H) ⁺ :328.1

Step 9: Synthesis of compound NJ-78

Compound 9 (150 mg, 0.5 mmol, 1.0 eq) was dissolved in DCM (10 mL), to which was added compound 2A (112 mg, 1.4 mmol, 3 eq), Cu(OAc) ₂ (183 mg, 0.91 mmol, 2 eq) and TEA ( 139mg, 1.37mmol, 3eq), reacted at room temperature for 36 hours. LC-MS detected that the reaction was complete, added disodium EDTA aqueous solution to the reaction system, stirred for 30 min, extracted three times with DCM, washed with brine once, and dried the organic phase with anhydrous Na ₂ SO ₄ . After drying, it was purified by silica gel column (PE/EA=15/1) to obtain the target compound NJ-78 (52.1 mg, 31.3%) as a yellow solid.

¹ H NMR (400MHz, CDCl ₃ ) δppm 8.70 (s, 1H), 8.61 (d, J=2.2Hz, 1H), 7.76 (dd, J=8.6, 2.2Hz, 1H), 7.65-7.59 (m, 2H) ),6.33(s,1H),2.67(s,3H),2.41(s,3H),1.42(s,9H).

LCMS: (M+H) ⁺ :364.1

HPLC: 99.43%

Example 2. Synthesis of compound NJ-95

The synthetic route of compound NJ-95 is shown in the following formula:

Experimental procedures and results for compound NJ-95:

Step 1: Synthesize 2

Sodium hydride (100 mg, 2.52 mmol, 1.2 eq) was dispersed in anhydrous tetrahydrofuran (15 mL), under nitrogen protection, cooled in an ice-water bath, and a solution of compound 1A (350 mg, 2.1 mmol, 1.0 eq) in tetrahydrofuran (2 mL) was added dropwise thereto. , and the reaction was completed at 0°C for 20 minutes. A solution of compound 1 (534 mg, 2.1 mmol, 1.0 eq) in tetrahydrofuran (3 mL) was added dropwise to the reaction solution, and the reaction was carried out at 0° C. for 40 minutes. The reaction was quenched with water and extracted with ethyl acetate. The organic phases were combined, washed with brine, dried over anhydrous sodium sulfate, and spin-dried to obtain a crude product. The crude product was purified by silica gel column (petroleum ether/ethyl acetate=15/1) to obtain the target compound 2 (700 mg, 87%, white solid). LCMS: (M+H)+: 384.2.

Step 2: Synthesis 3

Compound 2 (0.7 g, 1.82 mmol, 1.0 eq) was dissolved in ethanol (4 mL), an aqueous solution (2 mL) of potassium hydroxide (3 g, 54.8 mmol, 30 eq) was added thereto, and the mixture was refluxed at 85°C overnight. After the completion of the reaction was detected by TLC and LCMS, the ethanol was removed by rotary evaporation, and the pH value of the aqueous solution was adjusted to 6 with 2N dilute hydrochloric acid. The precipitated solid was filtered, washed with a small amount of water, and dried to obtain the target compound 3 (0.44 g, 64%, pale yellow solid).

1H NMR(400MHz, DMSO-d6)δ8.15(s,1H),7.42(d,J=8.3Hz,2H),7.37(d,J=8.3Hz,2H),2.55(s,6H),1.29 (s, 9H).

LCMS: (M+H)+: 375.1.

Step 3: Synthesis 4

Compound 3 (0.44g, 1.2mmol, 1.0eq) and DMF (one drop) were dissolved in dry dichloromethane (10mL), placed in an ice-water bath under nitrogen protection, oxalyl chloride (0.4mL, 4.8mmol, 4eq) was dissolved ) was added dropwise to the above solution, warmed to room temperature and stirred for 1 hour, spin-dried to remove the solvent and excess oxalyl chloride to obtain a crude aroyl chloride intermediate. This aroyl chloride intermediate was redissolved in dry dichloromethane (15 mL), placed in an ice-water bath under nitrogen protection, and aluminum trichloride (0.95 g, 7.17 mmol, 6 eq) was added to it in batches, and the addition was completed at room temperature. React overnight. Anhydrous ethanol (2 mL) was added to the reaction solution, and the reaction was carried out at room temperature for 1 hour. After the reaction was detected by LC-MS, dichloromethane (20 mL) and 1N dilute hydrochloric acid (15 mL) were added to separate the layers, and the aqueous phase was extracted with dichloromethane. (2×20 mL), the organic phases were combined, washed with brine (20 mL), dried, suction filtered, the filtrate was concentrated, and purified by column (petroleum ether/ethyl acetate=15/1) to obtain compound 4 (0.35 g, 76%), Pale yellow solid.

1HNMR(400MHz,CDCl3)δ8.94(s,1H),8.57(d,J=2.1Hz,1H),7.66(dd,J=8.4,2.2Hz,1H),7.48(d,J=8.4Hz, 1H), 4.39(q, J=7.1Hz, 2H), 3.17(s, 6H), 1.50–1.34(m, 12H).

LCMS:(M+H)+:385.1.

Step 4: Synthesize 5

Compound 4 (200mg, 0.52mmol, 1.0eq) was dissolved in methanol (3mL) and tetrahydrofuran (2mL), then sodium hydroxide (62mg, 1.56mmol, 3eq) aqueous solution (2mL) was added dropwise to the above solution, 50 ℃ React overnight. The organic phase was spun off, the aqueous phase was extracted with ethyl acetate (10 mL), the pH of the aqueous phase was adjusted to 3 with 2N dilute hydrochloric acid, the precipitated solid was filtered, washed with a small amount of water and dried to obtain compound 5 (110 mg, 59 %, pale yellow solid).

1HNMR (400MHz, DMSO-d6): δ13.34 (br s, 1H), 8.67 (s, 1H), 8.38 (d, J=1.8Hz, 1H), 7.83 (dd, J=8.4, 1.8Hz, 1H) ), 7.71(d, J=8.4Hz, 1H), 3.12(s, 6H), 1.36(s, 9H).

LCMS: (M+H)+: 357.1.

HPLC: 98.4%

Step 5: Synthesize 6

Compound 5 (8.2 g, 0.023 mol, 1.0 eq) was dissolved in acetonitrile (40 mL) and tert-butanol (40 mL), triethylamine (6.4 mL, 0.046 mol, 2.0 eq) was added, under nitrogen protection, and added dropwise to it DPPA (7.4 mL, 0.035 mol, 1.5 eq). After the addition was completed, the reaction was carried out at 100° C. for 5 hours. The reaction solution was concentrated and purified by silica gel column chromatography (DCM/MeOH=200/1) to obtain compound 6 (6.6 g, 67%) as a yellow solid.

¹ H NMR (400MHz, DMSO-d ₆ ) δ 8.72 (s, 1H), 8.52 (d, J=2.4Hz, 1H), 7.59 (dd, J=8.4, 2.4Hz, 1H), 7.44 (d, J=8.4Hz, 1H), 6.28(brs, 1H), 3.00(s, 6H), 1.47(s, 9H), 1.32(s, 9H).

LCMS: 428.2([M+H] ⁺ ).

Step 6: Synthesize 7

Compound 6 (6.6 g, 15.5 mmol, 1 eq) was dissolved in dichloromethane (20 mL), trifluoroacetic acid (10 mL) was added dropwise thereto, and the reaction was carried out at room temperature for 1.5 hours. The reaction solution was concentrated and extracted with aqueous sodium bicarbonate solution (100 mL) and dichloromethane (100 mL x 2). The combined organic phases were washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain the crude product. The crude product was purified by silica gel column chromatography (DCM/MeOH=100/1) to obtain compound 7 (3.9 g, 78%) as a yellow solid.

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.62 (d, J=2.0 Hz, 1H), 7.96 (s, 1H), 7.68 (dd, J=8.4, 2.0 Hz, 1H), 7.56 (d, J=8.4Hz, 1H), 3.71(br s, 2H), 3.06(s, 6H), 1.43(s, 9H).

LCMS: 328.2([M+H] ⁺ ).

Step 7: Synthesize 8

Compound 7 (2.5 g, 7.65 mmol, 1.0 eq) was dissolved in MeCN (30 mL) under nitrogen protection, to which was added CuI (1.74 g, 9.17 mmol, 1.2 eq) and t-BuONO (3.15 g, 30.58 mmol, 4.0 eq), react at 60°C overnight. LC-MS detected that the reaction was complete. After the acetonitrile was spun out, brine and ammonia water were mixed and added to the reaction system. Then DCM was added for extraction. 8 (1.0 g, 30%).

¹ H NMR (400 MHz, CDCl ₃ ) δ 8.97 (s, 1H), 8.50 (d, J=2.4 Hz, 1H), 7.60 (dd, J=8.4, 2.4 Hz, 1H), 7.43 (d, J= 8.4Hz, 1H), 3.16(s, 6H), 1.33(s, 9H).

LCMS: 439.0([M+H] ⁺ ).

Step 8: Synthesize 9

Compound 8 (750 mg, 1.71 mmol, 1.0 eq) was dissolved in dry dioxane (10 mL) under nitrogen protection, to which was added B ₂ Pin ₂ (2.17 g, 8.55 mmol, 5.0 eq), KOAc (340 mg, 3.42 mmol, 2.0eq) and Pd(dppf)Cl ₂ (130mg, 0.17mmol, 0.1eq), react at 90°C for 15 hours. LC-MS detected the reaction was complete, after cooling, concentrated to obtain crude product. Extract with water and dichloromethane (50 mL x 3). The combined organic phases were washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, and spin-dried to obtain the crude compound. The crude compound was purified with silica gel column (petroleum ether/EtOAc=20/1) to give crude compound 9 (200 mg) as a yellow solid.

LCMS: 439.2([M+H] ⁺ ).

Step 9: Synthesize 10

Compound 9 (0.42 g, crude) was added to THF (10 mL) and H ₂ O (4 mL) to dissolve, then NaIO ₄ (2.05 g, 9.6 mmol, 10.0 eq) was added, and the reaction was carried out at 50 °C for 15 h. Extracted with DCM three times and washed with brine once, the organic phase was dried over anhydrous Na ₂ SO ₄ and spin-dried to obtain crude compound 10 (250 mg) as a yellow solid.

LCMS: 357.1([M+H] ⁺ ).

Step 10: Synthesis of NJ-95

Compound 10 (80 mg, 0.2 mmol, 1.0 eq) was dissolved in DMSO (10 mL), to which was added compound 2A (83 mg, 1.2 mmol, 5 eq), Cu(OAc) ₂ (97 mg, 0.4 mmol, 2 eq) and TEA ( 74mg, 0.7 mmol, 3eq), reacted at room temperature for 120 hours. LC-MS detected that the reaction was complete, added disodium EDTA aqueous solution to the reaction system, stirred for 30 min, extracted three times with DCM, washed with brine once, and dried the organic phase with anhydrous Na ₂ SO ₄ . After drying, it was purified by silica gel column (DCM/MeOH=15/1) to obtain the target compound NJ-95 (6.9 mg, 8.0%) as a yellow solid.

¹ H NMR (400MHz, CDCl ₃ ) δ 8.70 (s, 1H), 8.61 (d, J=2.2Hz, 1H), 8.32 (s, 1H), 7.80 (dd, J=8.6, 2.2Hz, 1H) ,7.63(d,J=8.6Hz,1H),7.52(s,1H),7.27(s,1H),2.58(s,3H),1.42(s,9H).

LCMS: (M+H) ⁺ :350.0

HPLC: 96.05%

Example 3

Experimental materials and equipment: PC3 cell line (

CRL-1435 ^TM ), CCK8 (cat#CKO4, colleagues)

Experimental steps:

1. The inhibitory effect of each compound on PC3 cell proliferation was detected by CCK8 method. The cells in the logarithmic growth phase were seeded in a 96-well plate at a density of 1000 cells/well, in DMEM medium containing 10% serum, and continued to culture for 24 hours in an incubator at 37°C and 5% CO ₂ . Various compounds at different concentrations (0.2, 0.5 and 1 μM) were added, and a DMSO control group was set up, and each group was set up with 3 replicate wells. Complete medium containing 10% serum was used for drugs and cells. After co-cultivation for 96 hours in an incubator at 37°C and 5% CO2, the medium in the 96-well plate was gently shaken off, and 100ul of complete medium was added to each well. , and add CCK8 according to the standard of 10ul CCK8 reagent per 100ul medium, continue to incubate for 1 h in a 37°C 5% CO2 incubator, and measure the absorbance A of each well at a wavelength of 450nm. Cell viability=(average of drug group A/control group A)*100%

The results are shown in Figure 1. Different concentrations of NJ-78 and NJ-95 had no obvious killing effect on cell viability.

2. Biological function verification experiment of the compound: tumor cell migration experiment

PC3 cells were plated in a 6-well plate at a density of 5,000 cells per well, and an experimental group and a control group were established. The control group was added an equal proportion of DMSO, and the complete medium containing 10% serum was used to pre-culture in an incubator at 37°C and 5% CO2 for 96 hours, and then starved (with serum-free DMEM medium). After 24 hours to exclude the influence of cell growth, each well was digested into single cells, and resuspended in serum-free DMEM medium to form a single cell suspension with a density of 5*10 ^5. Continue to set up the experimental group and the control group, the experimental group All the medium (including the serum-free medium in the chamber) were added with 0.5, 1 and 2.5 μM of NJ-78 and NJ-95, respectively, and the control group was added with an equal proportion of DMSO. 500ul of 10% FBS was added to the Transwell wells. DMEM medium, and add 100ul cell suspension to each transewell chamber. Culture at 37°C, remove the chamber after 24 hours, discard the medium in the chamber, and fix it in 4% paraformaldehyde for 15min. Rinse with PBS several times , use a cotton swab to gently wipe off the possible remaining cells in the inner layer of the chamber, and place the chamber in crystal violet for staining for 20 minutes. Rinse with PBS for several times, and gently wipe off the excess crystal violet dye and PBS with a cotton swab. After the chamber is dry , observe and record the number of cells under the microscope.

The results are shown in Figure 2-3. After adding NJ-78 and NJ-95, the number of cells that migrated to the lower layer of the chamber decreased, and the migration ability of the cells decreased.

Among them, **** means p<0.0001, *** means p<0.001, ** means p<0.01, * means p<0.05, and the experimental results are expressed as mean ± S.E.M.

Specific embodiments of the present invention have been described above. It should be understood that the present invention is not limited to the above-mentioned specific embodiments, and those skilled in the art can make various changes or modifications within the scope of the claims, which do not affect the essential content of the present invention. The embodiments of the present application and features in the embodiments may be combined with each other arbitrarily, provided that there is no conflict.

Claims (9)

1. A class of compounds whose structural formula is as follows:

   

   Wherein, R ₁ is selected from lower branched alkyl, and R ₂ , R ₃ and R ₄ are selected from lower alkyl or H, respectively.
2. The compound according to claim 1, wherein the lower branched chain alkyl group is a 3-8 carbon alkyl group, and the lower alkyl group is a 1-8 carbon alkyl group.
3. The compound according to claim 1, wherein the structural formula of the compound comprises:

   
4. Use of a class of compounds as claimed in claim 1 in the preparation of a medicament for inhibiting migration of prostate cancer cells.
5. The use according to claim 4, wherein in the drug for inhibiting the migration of prostate cancer cells, the effective concentration of the compound is 0.5 μM-2.5 μM.
6. A pharmaceutical composition for inhibiting the migration of prostate cancer cells, characterized in that the pharmaceutical composition uses the compound according to claim 1 as an active ingredient.
7. The pharmaceutical composition for inhibiting migration of prostate cancer cells according to claim 6, wherein the total effective concentration of the compound in the pharmaceutical composition is 0.5 μM-2.5 μM.
8. The pharmaceutical composition for inhibiting migration of prostate cancer cells according to claim 6, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier or excipient.
9. The pharmaceutical composition for inhibiting migration of prostate cancer cells according to claim 6, wherein the composition comprises

   

   

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